DD222029A5 - METHOD FOR PRODUCING SUBSTITUTED VINYLCEPHALOSPORIN COMPOUNDS - Google Patents

Abstract

The invention relates to a process for the preparation of substituted Vinylcephalosporinverbindungen. The aim of the invention is to provide a method of the type mentioned, which leads to new and pharmaceutically useful compounds. The process according to the invention allows the preparation of new compounds which prove to be effective antibiotics and which can be used for the treatment of bacterial infections in medicine.

Description

15

Applicatio gsgobi et the f He indung

The invention relates to a process for the preparation of substituted Vinylcephalosporinverbindungen. The compounds obtainable according to the invention are useful in medicine for controlling bacterial infections. , .-

Characteristic of the known technical solutions

20 25

One possibility for preparing the 3-substituted vinylcephalosporins according to the invention consists in the use of 3-formylceph-3-em-compounds as intermediates: These intermediates can be obtained by oxidation of the corresponding 3-hydroxymethylceph-3-emes obtained by enzymatic hydrolysis the corresponding cephalosporins are available. This process is described in US Pat. No. 3,351,596, which discloses inter alia compounds II and III.

RCONH

CHO

M / 2 4 .2 79

  χ-

DI i (J

CH-

NH.

II

III

Derivatives of the 3-CHO group with carbonyl reagents, such as semicarbazide and. Hydroxylamine, disclosed, however, an alkylation of the carbon of the 3-CHO group is not described.

The corresponding sulfoxides are more stable and can be prepared in better yield (GB-PS 1 341 712).

3-Alkenyl-substituted cephalosporins were first described in QB-PS 1,342,241 (corresponding to U.S. Patents 3,769,277 and 3,994,884). Compounds IV and V can be found on pages 25 and 29 of the British Patent.

LJ

CHCONB I NH ₂

IV

CH ^ CONH

CH = CH.

H = CHCH.

M / 24 279

-Jf-

These compounds were riuich implementation of the corresponding 3-Tr i phenyl phosphor! iininine cephalosporins produced with formaldehyde or acetaldehyde. The inverse process, namely the reaction of a Phosphoranylidinderivate the. formula

10

15 20 '

30

RP = CR ³ H ⁴

with a 3-CHO-Cephalosporiπ is also described in this patent on page 5. It is mentioned in US Pat. No. 4,107,431 that compound IV is absorbed after oral administration.

Further, such compounds have been described by Webber et al. J. Med. Chem. IJi (IO) 986-992, (1975) and in US Pat. No. 4,065,620, which in columns 3, 4 and 5 indicate the type of compounds which comprise the compounds of the invention belong. Specifically disclosed were the compounds of formula VI:

CH = CHCO ₂ H

(CO ₂ Et) (CN)

VI

Further variations of this type are described in US Pat. Nos. 4,094,978 and 4,112,087, wherein compounds VII and VIII are listed.

35

M / 2 4 279

• -ET- 7

U.S. Patent 4,094,978 _; Column 44

CH = CHCH OH

HO

-CHCONH _ ..

NH ^ '/

2

-CH = CHCH ₂ OCH ₃ (trans)

VIII

U.S. Patent 4,112,087, column 31

Other 3-alkenyl-substituted cephalosporins are disclosed in the following publications;

U.S. Patent 3,830,700: 3- {nitrostyryl) cephalexin analogs

U.S. Patent 3,983,113 U.S. Patent 4,040,806 U.S. Patent 4,139,618;

U.S. Patent 4,147,863:

3- {heterocyclothio) propenylcephalosporins

\ 3- (1-methyl-5-tetrazolyl) vinyl

cephalosporins. '

M / 2 4 2

-AT- f)

DK-OS 30 19 445: 3- (sulfonyloxy) vinylcephalosporins

FR-PS 24 60 302: 3- (dimethyl on J no) vinylcephalexin

analo-ga

EU PS 30 630:

U.S. Patent 4,255 U.S. Patent 4,390,693:

7-1 (3-Met hansulί onamidophenyl) -aminoacetamido] -S-vi 4-carboxylic acid

7- (2-Thienyl) acetamido-3- (3-acetoxy-1-propenyl) and -3- (heterocyclovinyl) ceph-3-em-4-carboxylic acids and 7a-methoxy analogs.

The most important commercially available cephalosporins - the compounds according to the invention are intended for the same purpose - are cephalexin, cefadroxil, cephradine and cefaclor. These compounds correspond to the formulas IX, X, XI and XII.

CHCONH I

NH ₂

30 R = H Cephalexin I R = OH Ceradroxil X.

CO ₂ H

CHCONH

Cefaclor XI

Cephradin XH

M / 2 4 28 0. - ^ j

These compounds are the subject of the following patents:

Cephalexin U.S. Patent 3,507,861

Cefadroxil -US-PS 3 489 752 '

5 (Re 29,164)

Cefachlor -US-PS 3 925 372 \,

Cephradine -US-PS 3 485 819

Compounds of similar structure are 3-chlorocefa-O droxin and 3-hydroxycefadroxil described in U.S. Patent 3,489,751 and British Patent 1,472,174.

All known compounds have disadvantages in terms. their effectiveness and applicability.

'' Aim of the invention

The aim of the invention was to provide new cephalosporin compounds, which are superior to the known compounds in terms of effectiveness and applicability.

Explanation of the essence of the invention

The object of the invention was to provide a process for the preparation of such cephalosporin compounds. This object is achieved by the process according to the invention for the preparation of substituted vinyl cephalosporin.

compounds of general formula XV 30

(XV)

i J I

M / 24 280

-JfC- r

wherein

η is 0 or 1,

R is a hydrogen atom, OP is a lower alkoxy group

or a halogen atom, P, P and P denote hydrogen atoms or customary protecting groups for amino-, carboxy- and hydroxy groups used in cephalosporin chemistry,

2 3

R represents a hydrogen atom, OP or a lower alkoxy group, and

R is a hydrogen atom, a C -C alkyl,

C ₇ -C ₁₄ aralkyl, heterocyclothio, C 1 -C 4 alkyl, C 1 -C 4 alkoxy-C 1 -C 4 alkyl group or the group AIkX, where

Alk is an alkylidene or alkylene radical having 1 to 4 carbon atoms, and

X represents a bromine, chlorine or iodine atom,

and

the pharmaceutically acceptable acid addition salts,

Metal salts, amine salts or esters thereof.

Preferably, the process is for the preparation of the compounds of formula XV, wherein R is CH_ or AIkX, to obtain the compounds of formulas XIII and XIV:

CH = CHCH.

M / 2'l 27 9

CH = CHAIkX

XIV

in which

η is 0 or 1,

1 '3

R represents a hydrogen atom, OP, a lower alkoxy group or a halogen atom, namely a chlorine, bromine, fluorine or iodine atom,

12 3

20 ρ, ρ and P are hydrogen atoms or in

cephalosporin chemistry commonly used protecting groups for amino, carboxy and hydroxy groups ,.

R represents a hydrogen atom, OP or a lower alkoxy group,

Alk represents an alkylidene or alkylene group having 1 to 4 carbon atoms, and

X represents a chlorine, bromine or iodine atom.

M / 24 279 ·. -y

In the context of the present invention, the term "lower" in connection with alkyl, alkoxy groups and the like means a hydrocarbon radical having 1-8 carbon atoms, preferably '1-6, most preferably 1-4 carbon atoms.

Those compounds of the above formulas in which. ^{n is} 1 and P, P and P - represent conventional protective groups are intermediates for the preparation of the biologically active end products according to the invention which are described by the general formula XIII,

12 3

wherein η is Q, and P, P and P are hydrogen atoms. These compounds are useful as orally effective cephalosporin antibiotics having potent activity against Gram-positive bacteria. They also have an improved spectrum of activity compared to cephalexin, cefadroxil, cefaclor and cephradine against Gram-negative bacteria, difficult to control bacteria and anaerobes. Further, after oral administration, they result in a prolonged circulating antibiotic concentration and are suitable for administration to humans based on one or two doses per day. As such, they are administered in doses of .100 to 5,000 mg per day, depending on the size of the patient and the disease state. They may also be administered parenterally in similar dosages.

The compounds of formula XIV are of interest mainly as intermediates. Those connections,

12 3

However, where η is 0 and P, P and P are hydrogen, they have antibacterial activity and are also useful as antibiotics.

M / 24 279. -10

In view of this property, the compounds of the formulas XIII and XIV in which η is 0 and P, P and P represent a hydrogen atom are for the treatment of bacterial infections caused by sensitive organisms; useful in mammals, including humans. For this purpose they are administered orally or parenterally in an antibacterially active, non-toxic dose as such or in the form of a pharmaceutically acceptable acid addition salt, pharmaceutically-acceptable metal or amine salt or a pharmaceutically acceptable ester.

Pharmaceutically acceptable acid addition salts are those in which the anion does not significantly contribute to the toxicity of the salt and which are compatible with the usual pharmaceutical carriers and suitable for oral or parenteral administration. Such salts

The salts of compounds of the formulas XIII /. and XIV in which η is 0 and P is hydrogen atom, with mineral acids such as hydrochloric acid, hydrobromic acid, phosphoric acid and sulfuric acid, with organic carboxylic acids or organic sulfonic acids such as acetic acid, citric acid, maleic acid, succinic acid, benzoic acid, "tartaric acid, fumaric acid, Mandelic acid, ascorbic acid, malic acid, methanesulfonic acid, p-ToJ-Uolsulf onsäure, as well as with other, on penicillin and cephalosporin area known and used

OQ acids. The preparation of such salts is carried out by conventional methods by reacting one of the compounds of formulas XIII or XIV, wherein η is 0 and P is a hydrogen atom equivalent amount of the acid.

P represents a hydrogen atom, with a substantially

M / 24 279

Pharmaceutically acceptable metal and amine salts are those salts of the compounds of formulas XIII and XIV wherein η is 0 and P is hydrogen, which are stable under ambient conditions and in which the cation does not significantly contribute to the toxicity or biological activity of the salt , Suitable metal salts are sodium, potassium, barium, zinc and aluminum salts. Sodium or potassium salts are preferred. Amine salts prepared from, for example, benzylpenicillin-related amines and capable of forming stable salts with the acidic carboxyl group are trialkylamines such as tri-ethylamine. Procaine, dibenzylamine, N-benzyl-.beta.-phenethylamine, 1-ephenarnine, N, N- ^1- dibenzylethylenediamine, dehydropabietylamine, N-ethylpiperidine, benzylamine and dicyclohexylamine.

Pharmaceutically acceptable esters are those esters which are effective per se or which are pro-drugs in that they form in the body to form the antibiotic

be hydrolyzed per se. Suitable esters of the last type are phenacyl, acetoxymethyl, pivaloyloxymethyl-acetoxyethyl, a-acetoxybenzyl, a-pivaloyloxyethyl, 3-phthalidyl, 5-indanyl, methoxymethyl, benzoyloxymethyl, a-ethylbutyryloxymethyl, propionyloxymethyl , Valeryloxymethyl, isobutyryloxymethyl, glycyloxymethyl esters and other esters known in the penicillin and cephalosporin art.

The compounds of formulas XIII and XIV, wherein η is for

12 3 0- and P, P and. P represents hydrogen atoms, and their salts as defined above may be formulated for oral or parenteral use in a conventional manner using known pharmaceutical carriers and excipients

M / 24 279 -12-

become. They may be presented in unit dose form or in multi-dose containers. The pharmaceutical agents may be in the form of tablets, capsules, solutions, suspensions or emulsions. The compounds may also be used in the form of suppositories _; conventional suppository bases, such as cocoa butter or other fatty materials. If desired, the compounds may be administered in combination with other antibiotics, including cephalosporins, penicillins and aminoglycosides.

The above statements apply equally to the compounds of the formula XV.

Table 1 contains a summary of the structure. Features of those described in Examples 1 to 43

2Q connections. Most of these precursors are 7β- (D-phenylglycyl-inido) cephalosporins with a 1-propen-1-yl group in the 3-position. The terminal carbon atom of the propenyl group is substituted in some compounds, for example with an alkyl group

"Γ (methyl), a halogen atom (chlorine or iodine atom), an aryl group (phenyl group), a heterocyclothio group (1,2,3-triazol-5-yl-thio radical) or an alkoxy group (methoxy group). The phenylglycylamido group may be unsubstituted or substituted with a hydroxy, alkoxy or halo group.

oQ logengruppe mono- or disubstituted.

M / 24 279

table

Compounds of Examples 1-4 3

CH = CKR ^-

(XV)

Compound No. 9 (BMY-28100)

13 (BBS-.1058),

11 (BBS-1064)

24 (BBS-1065)

26 (BBS-10'66)

β (BBS-1Q67)

15 (EBS-1076)

21 (B3S-1091)

17 (BBS-1092!

32 (BMY-28060) 37 (BMY-28068)

Cl

HC R ^ R ^J (configuration) OH -CH, (Z)

(Z)

-H

K -CH ₃ (Z) K .Κ -CH ₂ Cl (Z) H ΟΚ -CH ₃ (E) K- OH - ^CH 2 ^C 6 H ₅ (Z) H OH -CH ₂ -S J "N J Ii I H

-CH ₂ OCH ₃ (Z)

(Z)

-CH ₃ (Zj

42 (BMY-28097)

CK.C

-CK ₃ (Z)

M / 24 279 -14-

Table 2 summarizes in vitro antibacterial activities of compounds of the invention. The activities are reported in terms of minimal inhibitory concentrations determined by the agar dilution technique for three groups of organisms designated Gp-Ia, Gp-Ib and Gn-Ia. Each of these organism groups is composed of five individual microorganism strains, which are indicated in the footnote to the table. The Gp-Ia organisms are Gram-positive staphylococci that are penicillin-sensitive. The Gp-Ib organisms are Gram-positive staphylococci that are penicillin-resistant and produce penicillinase. The Gn-Ia organisms are Gram-negative bacteria that are ampicillin and cephalothin sensitive. The compounds of the invention generally have low activity against ampicillin and cephalothin resistant Cram negative bacteria. From Table 2, with respect to the in vitro antibacterial activity of these compounds, the following conclusions can be drawn.

All compounds have good activity against penicillin-sensitive staphylococci (Gp-Ia). The activity against penicillin-resistant staphylococci (Gp-Ib) is generally lower by a factor of three or more. In any case, however, the compounds are many times more potent than cephalexin and cefadroxil.

Only those compounds which have an unsubstituted 3-position eis (Z) -propenyl group have good activity against Gram-negative bacteria (Gn-Ia). This is apparent from the compounds Nos. 9, 24, 32 and 42. The activity of the trans (E) -propenyl compound no. 8 is compared to the corresponding one

M / 24 279. -15-

cis-propenyl compound no. 9 lower by a factor of 8. Similarly, substitution at the terminal methyl group of the 3-propenyl substituent appears to result in reduced activity against Gram-negative bacteria, cf. Compounds Nos. 13, 15, 21 and 17. This also applies to the vinyl compound No. 11. Nevertheless, these compounds are potent antibacterial agents that are substantially equivalent to cephalexin and cefa- droxil. Ring substitution is in no way detrimental to antibacterial activity, cf. Compounds 9, 24, 32 and 42. Compound No. 37 appears to be an exception to the above conclusion conclusions, but as shown in Table 3, it is a potent active substance against both Gram-positive and Gram-negative bacteria ,

20 ·

Table 2

Agar dilution technique (Mut: 1 ler-Hinton agar) Minima le inhib concentration (mcg / ml)

(BMY-28100), • Gd Ia ³ 1 gc ib 3 gn 8th i-J .7 (BBS-1058) 1 ι 0th 2 1 1 1 Connection no. . (BBSt.1064) 12:23 12:35 1. S2 0th 8th .0 · 6 0 9 (BBS-1065) 12:40 1. 4 4th 8th .8th 13 (BBS-1067) 12:40 0th 2 Third 3 11 (B3S-1076) 12:23 0.3 1'. 92 0th 52 0th 0 24 (BBS-1091) ' 12:26 0th 4 6th 7 8th (BBS-1092) 12:20 Second 7 > 50 7 15 (BMY-2S060) 0.61 Second 7 Second .1 21 . (BMY-28068) 12:53 1 Second .3 17 C.12 0th 53 1 32 6:30 7th 2 6 37

M / 24 279 -IG-

Gp Ia ^J Gp Ib ³ Gn Ia ³

Connection No. 1 2_. 1_ 2 1 2

(BMY-28097) 0.35 1..2 * 0.53 *

cephalexin ^ 1-2 0.70 4.1-3.6 6.2

Cefadroxil 1.2 1.10 3.6 4.1 8.3

1 · Columns 1 and 2 mean separate tests.

2. Columns 1 and 2 mean separate tests.

3. Mean for five organisms per group:

Gp Ia Gram + Staphylococci; Penicillin-sensitive; no penicillinase producers: J5 S. aureus Smith A9537

S. aureus A 9497

S. aureus Terajima

S. aureus A9534

S. aureus A9601

Gp Ib Gram + Staphylococci; Funicillin-resistant;

Penicillinase producers: S. aureus 19 3 S. aureus BX-16J3-2 A96Ü6 25 S. aureus A15092

S. aureus proboscis] S. aureus A9602

Gn Ia Gram Bacteria; Ampic j 1.1 Ln- 'and Cephalothin-OQ sensitive: *

E. coil Juhl Al 51 IV

E. coli A9660

K. pneumoniae DIl

P. mirabilis A9554 _Q5 P. mirabilis A9900

M / 24 279 -17-

4. Not part of experiment 1; the values were obtained separately. , ,

  "

Table 3 contains Verglui chsdatien for the 'in vitro antibacterial activity against the same organisms, as arigegeben in Table 2, where two different bacteriological culture media were used. Miller-

IQ Hinton agar is the standard medium used in the tests described in Table 2. Table 3 contains a comparison of the minimum inhibitory concentrations of three of the test compounds, which are first determined in Mueller-Hinton's medium and then in nutrient agar

were jg. ¹ Compound No. 9, which is substituted at the 4-position of the phenyl ring by a hydroxy group, and the compound No. 42, which has a 3-methoxy-4-hydroxy substitution pattern on the phenyl ring, have only moderate medium ef ect. * This is meant to express

2Q that the MIC values differ by less than a factor of 3. The 3,4-dihydroxyphenyl-substituted Compound No. 37 gives a difference in activity between the two media by a factor of 6 to 12, with the minimum inhibitory concentrations determined in nutrient agar which are much lower than those determined in Mueller-Hinton agar. As a result, Compound No. 37 is comparable in its antibacterial effect to the other cephalosporins indicated in Table 2, which are in the 3-position

have gg cis-propenyl group. The phenomenon of different antibiotic activity in different nutrient media has been previously studied and published, cf. T.A. Pursiano et al. Antimicrobial Agents and Chemotherapy, Part 3, No. 1, pp. 33-39

M / 24 279 -1.8-

Tabelle._l

Comparison of the test media

Agar dilution technique

Minimal inhibitory concentrations (mcg / ml)

10 Connection no. (BMY-2810.0) A ¹ Gp Ia Gp Ib Gn Ia ² 9 B 12:23 0.92 0.70 (BMY-28068) A 12:17 12:35 0.70 15 37 B. 4.8 * 6.3 5.5 (BMY-28097) A 12:40 ν 0.61 0.92 42 B 12:35 1.2 12:53 12:23 12:40 12:40

A Muel ler-liiiiLüii agar B _:. Nutrient agar

2

, Mean values for the groups

Organisms as in Tube.l J t. · 2.

The structure-activity correlations derived from the preceding in vitro studies are confirmed by in vivo studies in mice. Table 4 summarizes the respective shot dose for mice that were infixed with a lethal bacterial infiltrate. Two different types of bacteria were used, namely a Gram-positive organism and a Gram-negative organism. The protective dose (PD, -) is the dose administered after administration

M / 24

a group of infi zicu do Mciusun leads to a survival of 50 nach after five days. Normally, untreated infected mice undergo treatment within three days after injection of the lethal inoculum.

Tab rc e 4

Protective dose 'for mice with a lethal inoculum

Oralo were administered

Connection no.

9 (BMY-28100)

13 (BBS-1058) 11 (BBS-1064) 24 (BBS-1065)

(BES-1067)

32 (BMY-28060)

S. aureu Smith '0.14 (0.31) 0.32 (0.31) 0.18 (0.31) 0.18 (0.27) 0.20 (0.31) 0.17 (0.22)

E. coli Juhl

1.2 (8.4)

3.0, (8.4)

3.8 (8.4)

1.5 (8.2)

7.5 (8.2)

3.04 (8.4)

37 (BMY-28068) 42 (BMY-28097)

0.13 (0.27) 0.09 ³

0.44 (8.2)

M / 24 279 -20-

Dose in mg / kg, the 5 days the death of 50% of the animals in groups of 5 mice, which with verschiep. which doses of the compounds to be tested were treated on the day of infection prevented; the values were determined by interpolation from the dose-response curve; untreated animals die within 3 days.

10 2

The values in brackets were in the same experiment '

obtained for cephalexin.

In this experiment, a value of 0.16 mg / kg was obtained for ÜMY-28100; Control values for cepha- «

^λ .

lexin or cefadroxil are not available.

The data in Table 4 were obtained from several different experiments. Cephalexin was used as reference compound, the PD ^ values for this compound in the same experiment are given in parentheses next to the PD, ₀ values of the compounds to be tested. It can be seen that all cephalosporins have a good activity against Gram-positive Staphylococcus aureus infections and that the compounds which have a cis-propenyl group in the 3-position are more effective against infection with Gram-negative bacteria, cf. Compounds 9, 24 and 37.

Table 5 contains comparative blood level data 30

for mice treated orally and intramuscularly with the compounds listed in Table 1. It can be seen that with the exception of compound no. 21, which has a heterocyclothio substituent on the 3-propenyl group, all compounds are equally well orally, '.

be absorbed. The compound no. 37 gives

M / 24 279 -21-

  '

oral administration exceptionally high blood levels in mice. It has been shown that this compound is metabolized in rats in Compound No. 4 2, cf. Example 43. Compound No. 37 is the 3,4-dihydroxyphenyl compound while Compound No. 42 is the corresponding 3-methoxy-4-hydroxyphenyl compound which has strong in vitro and in vivo activity.

,ο, ''

25 ·

10

Table 5 Blood Levels in Mice Dose 100 mgAgr po . 20 mg / kg ι Ρ · ⁰ ·

20

Connection no. C max (mcg / ml) Hours. ÄUC (meg std / ml Trial 1 9 (BMY-28100) 56 1.9 106 (lot 2) 13 (BBS-1058) 51 1.9 150 11 (BBS-1064) 43 1.2 49 24 (BBS-1065) 40 1.1 84 8 (BBS-1067) 30 1.4 69 15 (BBS-1076) 41 2.7 81 21 (BBS-1091) 4.4 3.2 19 17 (BBS-1092) 73 1.7 197 Cephalexin * 47 1.4 57 Cefadroxil 56 2.3 103 Trial 2 9 (BMY-28100) 61 1.3 86 24 (BBS-1065) 33 1.1 46 32 (BMY-28060) 25 1.7 37 37 (BMY-2806B) 180 2.5 666 Cefadroxil 51 1.5 67

max 1/2 "" ^ ^ max * l / 2

Std . (megs hours / ml (mcg / ml (hr ) (meg std / ml) (mcg / ml) (hrs) (meg, hrs / ml)

* Mean of 3 trials

1.9

1.6

26

13 2.0 41 11 1.4 13 7.8 1.3 15 10 ^- 1.3 22 12 2.7 3B 1.7 2.6 6.6 18 1.6 2 B 11 1.3 14 12 1.2 18

15 .5 1.7 13 9 .9 0.69 13 7 1.7 13 58 5.1 270

21

28

0.!

25 0.74 .20 23 0-37 15 31 12:50 -22 31 0.63 3! 31 0.99 52 16 0.5Ί o.f, 2Ί n.fin 19 26 Ο.Ί0 21 12:33 IA

21 0.48 13

16 0.58 13

21 0.48 13

86 1.2 233

30 0.37 2L

25

M / 24 279 -23-

Table 6 contains further in vivo data for Compound No. 9 versus four other organisms compared to cephalexin, cefachlor and cefadroxil. Tables 7 and 8 contain in vitro comparative data for Compound No. 9 versus cephalexin, cefadroxil and cefachlor for a variety of Streptococci, Neisseria, Haemophilis and various egg shakers.

10. ,

In attempts to recover the compounds from rat urine, it has been found that the value for the recovery of compound No. 9 from the urine is treated orally.

i '

after 24 hours with cephalexin and

.., - cefadroxil value is comparable and greater than the value obtained for cefachlor. Stability studies in which Compound No. 9 with cephalexin and cefachlor in solution using phosphate buffer at pH 6.5 and pH 7.0, human serum (pH 6.8), equine

₀ serum (pH 7.6) and calf serum (pH 8.2) as carriers, have demonstrated that the stability of compound no. 9 is greater than that of cephachlor and comparable to the stability of cephalexin.

table

Protective dose PD, ₀ in mice treated with a

lethal inoculum were infected with oral treatment

Organism S. aureus BX-1633

9 (BMY-28100) cephalexin

2,2 -

Cefachlor 2,2

Cefadroxil

7.2

S. pyogenes A20201

Hi

influenza A9 72 9

P. mirabilis A9554

0.11 1.8

0.74

18 0,

1.6 1.8

0.25

14

to CN

co

Table 7 Irf Vitro activity against Streptococci, Neisseria and Haemophilis

MIC (mcg / ml)

organism * A9604 A9505 9 (BMY 15 (BBS Cephalexin Cefadroxil cefaclor "A20065 A15069. 28100) 1076) S. pyogenes. S-23 ¹¹ A15040 Geometric mean (Lot 2) , 0.2 0.0 0.0 Thick Geo-metrical N. gonorrheae 'A15112 0.2 0.0 0.0 0.2 S. pneumoniae type II "A20142 0.05 0.2 0.0 0,0 _ 0.2 "Type I A20143 0.05 0.2 0.0 0.0 0.2 Type IH A20154 0.05 0.2 0.0 0.0 0.2 "A20155 0.05 0.20 0.00 0.00 0.2 Geometric means 0.05 0.2 3.1 1.6 • 0.2 0,050 0.2 3,1 ' 1.6 1.6 0.2 • 0.2 3, .l 1.6 1.6 0.2 0.4 3.1 1.6 0.2 0.2 3.1 1.6 i, c 0.2 0.23 3.1 1.6 0.8 0.2 > 100 6.3 6.3 1.3 · 0.20. > 100 6.3 6.3 0.8 0.0 > 100 6.3 6.3 0.0 > 100 6.3 6.3 0.8 ν 0,0 > 1OO 6.3 6.3 0.8 0.0 MOO 6.3 6.3 0.8 0.0 0.8 α, oo

co

(Jl

ω O

to O cn

Table 7 (continued) Organism

N. meningiticlis

A20048 A2004 9

9 (BMY-28100) IiQt 2)

0.8

15 (BBS_I0Z6.L_

> 100 cephalexin cefadroxil cefachlor

6.3

M A21487 A9729 A22157 0.8 > 100 6.3 Il A21496 A20177 A22481 0.8 > 100 6.3 Il A21497 A20193 A22491 0.8 ^: > 100 6.3 Geometric mean A21523 A20201 0.80 > 100 6.3 H , influenzae A9833 A20759 0.8 > 100 6.3 Il A22483 0.8 > 100 6.3 " A22482 " 0.8 > 100 6.3 Il Geometric mean 0.8 > 100  6 ,? " , influenzae 0.8 > 100 6. '; 2 Il Il 0.8 > 100 , -6.3 II 11 0.8 > 100 6.3 • pyogenes 0.80 , > 100 6.3 H pneumoniae 1.6 25 3.1 1.6 .25 3.1 f>, 6 25 3.1 S 0.1 0.2 0.8 S 0.2 0.4 3.1

6.3

6.3 6.3 6.3 6.3

6.3 6.3 6.3 6.3 6.3 6.3 6.3 6.3 6.3 6.3 6.3 6.3 0.4 3.1

0.8 0.8

0.8 0.8 0.8 0.8

0.8

ο, έ

0.8 0.8 0.8 0.4 0.4 0.66

cn

CO O

FCO

cn

O cn

Table 8 In Vitro Activity vs Anaerobes

cn

organism

ß-lactamase

MIC (mcg / ml)

9 (BMY-28100)

Cephalexin cefachlor

Gn, chopsticks B. fragilis

A20928-1 A21900 A20935 Geometric mean

0.8 50 0.8 3.2

12.5

12.5

6.3

9.9

3.1 6.3 3.1 3.9

Gn, chopsticks B. fragilis

A22053 A22021 A22693 Geometric means

Moo moo moo

25

100

-> ioo

> 75

100> 100

Moo> 100

A22695 A22533 Geometric mean

> 100 MOO> 100

100> 100> 100

100> 100,> 100

Gp, chopsticks Gp. Cocci

C. difficile A21675

C. perfringens A9645

P. acnes A21933

P. anaerobius A21905

Geometric mean

6.3 0.4 0.4 0.8 0.95

100 12.5 1.6 6.3 11

25 1.6 0.8 0.4

♦ clindamycin-resistant

M / 24 -279

The preparation of the environmentally useful compounds is accomplished by the synthetic routes described in GB-PS 1 342 241, US Pat. Nos. 3,994,884 and 4,107,431, mentioned above in connection with the appropriate starting materials. Substitution of the substituted vinyl group at the 3-position of the compounds of this invention essentially involves reacting a halide compound with a triarylphosphine to form a phosphonium salt which is converted to a phosphoranyl compound by treatment with a base. This is then reacted with a carbonyl compound to the compounds of the invention. In this case, either the halide compound or the carbonyl compound contains the cephalosporin core. The course of the reaction is illustrated by the following reaction schemes.

halide

QCH ₂ X

Base Reaction Scheme 1

Reaction scheme 2

Phosphoranyl compound

[QCH = P (C ₆ H ₅

carbonyl

C = O

QCH = CHR ^-

CHX (C.H.) - P

o2?

base

C = P

QCH = O

"QCH = CH R deblocking

      , - 'Acylation -

ii) I) In the case of XIII or XIV, XV

M / 24 279

In these reaction schemes, R represents a hydrogen atom, a C 1 -C 6 alkyl, C 2 -C 4 alkoxyC-C ₁ -C ₄ alkyl, C ₇ -C ₁ aralkyl, or AlkX group, wherein Alk and X have the meanings given above. Q is the 7-amino-3-cephem-3-yl-4-carboxylic acid nucleus, wherein the amino and carboxylic acid groups may have protecting groups, such as silyl groups or other groups known to those skilled in the art of beta-lactam antibiotics, or a 7-acylamino-3-cephem-3-yl-4-carboxylic acid moiety, wherein the 7-acylamino group represents a group which is more commonly

\

is present in cephalosporin antibiotics, including the invention used in the form in XiII and IX defined a-amino-a-substituted phenylacetamido group. The sulfoxides of the compounds mentioned have advantages. More specifically, Q represents a group of the following formulas:

(0)

-CCONH NHP '

(P)

(0)

AcNH

M / 24 279 '-30-

wherein

5 R has the meanings given above,

η is 0 or 1 and relates to the number of oxygen atoms bound to the sulfur,

Ac is an acyl group commonly present in T-acylaminocephalosporins, such as phenylacetyl or phenoxyacetyl, and

B denotes an aldehyde or ketone-derived alkylidene or aralkylidene protecting group, such as the benzylidene group, which is readily cleaved in a subsequent stage of the reaction, for example by hydrolysis using Girard's reagent T, and P / P _? and P represents hydrogen atoms or protecting groups commonly used in cephalosporin chemistry for amino, hydroxy and carboxyl groups.

2 Suitable carbonyl protecting groups (P) are, for example, aralkyl groups such as benzyl, p-methoxybenzyl, p-nitrobenzyl and diphenylmethyl (benzhydryl) alkyl groups; Alkyl groups such as the t -butyl group; Haloalkyl groups such as the 2,2,2-trichloroethyl group and others described in the literature, for example in GB-PS 1 399 086.

gO bene carboxyl protecting groups. It is preferable to use carboxyl-protecting groups which are easily removed by treatment with acids, in particular the benzhydryl or t-butyl group.

, M / 24 279 -.31-

, ,

Amino and hydroxy protecting groups (P and P) are known to those skilled in the art and include, for example, the trityl and acyl groups, such as the chloroacetyl, formyl, trichloroethoxycarbonyl, t-butoxycarbonyl or carbobenzyloxy group, etc. Again, amino protecting groups are preferred which are readily available can be removed by treatment with acid, in particular the t-butoxycarbonyl group.

If the cephalosporin core Q in the reaction schemes

1 and 2 in the form of 1-oxide (n = l) is used, these oxides are prepared by known methods, such as oxidation of the corresponding cephalosporin (n = 0), with ι-m-chloroperbenzoic acid or peracetic acid ago. In a later stage of the synthesis, the 1-oxide is reduced by known methods, for example with iodide ions in an aqueous medium.

The conversion of the halide compound of the formula QCH-X according to Scheme 1 into the phosphoranyl compound is preferably carried out using a halide in which X represents an iodine atom. If a chloride or bromide is used as the halide compound, it can first be converted to the corresponding iodide by treatment with sodium iodide in dimethylformamide or acetone. The iodide compound readily reacts with a triarylphosphine, such as triphenylphosphine, in an organic liquid carrier which is opposite to the

oQ reactants is inert under the reaction conditions.

Suitable conditions are room temperature and a reaction time which can be short, but which can also extend over several hours. In addition to triphenylphosphine, suitable triarylphosphines are such compounds

gc which are readily available and have reactive aryl groups such as substituted phenyl

M / 24 279 -32-

groups, e.g. Tolyl, naphthyl, substituted naphthyl and heteroaromatic or substituted heteroaromatic groups. The first reaction step involves the formation of the trxaryl phosphonium salt, which usually precipitates out of solution and is filtered off. The triarylphosphonium salt is then dissolved in a suitable liquid organic solvent which is immiscible with water and inert under the reaction conditions.

i. '

For example, in chloroform, trichlorethylene or other polychlorinated or -bromierten methane or ethane compounds. The phosphoranyl compound is then prepared in situ by treating this solution with aqueous alkali metal carbonate bicarbonate, or hydroxide at room temperature. The organic layer containing the phos- phoranylverbindung is separated, washed with water ^and dried in the usual way. Subsequently, one adds to the dry solution of the phosphoranyl compound the Carbonylverbinduhg listed in the reaction scheme, the last reaction step then takes place at room temperature in a relatively short time or for about 2 to

20 hours. The desired product of the formula

3 '

QCH = CHR is determined by known laboratory methods, such as

25 matography on a silica gel column.

The halide compounds of the formula QCH-X given in scheme 1 are obtained from the corresponding 7-amino- or 7-acylamino-3-hydroxymethyl-ceph-3-em-4-carboxylic acid derivatives by methods known in principle.

The conversion of the halide compound of the formula R CH X into the phosphoranyl compound according to Scheme 2 can be carried out with the corresponding chloride, bromide or iodide compound (X = Cl, Br or J). If desired, the chloride or bromide compound can be prepared as described above.

M / 24 279 -33-

However, this step is not essential. The reaction with the triarylphosphine, such as triphenylphosphine, is carried out either without a solvent or in an organic liquid carrier which is inert under the reaction conditions. The reaction can be carried out at room temperature or elevated temperatures for 1 to 24 hours at 20 to 150 ^° C. The triarylphosphonium salt usually precipitates and is filtered off. It is then dissolved in a suitable liquid organic solvent such as dimethyl sulfoxide or a water immiscible solvent such as ether, or tetrahydrofuran, and with a base such as butyl lithium, phenyllithium, sodium methoxide or sodium hydride for a period of several minutes to treated at a temperature in the range of -40 ⁰ C to +50 ⁰ C for a few hours. Are then added to the dry reaction solution, the carbonyl compound and allowed to react one to several hours at -40 ° C to +50 ⁰ C. The desired product of the formula

H QCH = C ^

^ R ^3:

is obtained as described above.

The reaction sequence according to Scheme I is particularly suitable ^{for the} preparation of those compounds of the formula

QCH = CHR, wherein R is lower alkyl, phenylalkyl, naphthalkyl, haloalkyl or alkoxyalkyl with cis (Z) -konfi-. guration stands. In one method, referred to as A variation of the reaction sequence according to Scheme 1 .g§ 7ß- [is - [a (N-1 -but carbonyl oxy- 1 ami no) -a- (ρ-hydroxyphenyl) acetamido] -B-chloromethyl- B-cephem ^ -carboxylic acid as

M / 24 279 -34-

1

Halide compound used. This method will be explained with reference to Examples 4, 5 and b.

Another variation of the reaction sequence according to Scheme 1, which is similar to process A, consists in using 7β- [α- (N-1-Bu toxycarbonylamine in ω) -α- (ρ-hydroxyphenyl) -acetamido] -B Chloromethyl-S-cephem ^ -carboxylic acid benzhydryl ester is used as the starting material, but in this reaction, referred to as Process B, chloroacetaldehyde is used to prepare the blocked 7-aminocephalosporanic acid having the 3-chloro-1-propene-1-yl group in 3 position produce. This latter compound has antibacterial activity, although not excellent. In Method B, the 3-chloro-1-propen-1-yl compound is used as an intermediate and first converted to the corresponding 3-iodo-1-propene-1-yl compound, which in turn is then added with a heteroaromatic thiol the 3-Heteroarylthioprop-l-en-l-yl-cephalosporin derivatives is reacted.

Another variation of the reaction sequence according to the scheme is called method C. As described above, the 7-amino-3-chloromethyl-3-cephem-4-carboxylic acid benzhydryl ester is prepared and the 7-amino group is protected by reaction with benzaldehyde to give the benzylidene protective group.

The latter compound is then converted to the phosphonium salt by treatment with triphenylphosphine, which is subsequently converted with a base to the corresponding phosphoranyl compound. This is treated with an aldehyde, whereby the 3-substituted vinyl

OR 7-aminocephalosporanic acid, which can then be acylated to introduce the desired acyl group in the 7-position.

M / 24 279. -35-

From the reaction sequence according to Scheme 2, 2 variations are proposed. According to the first, method D, as described above, ^{1 is} prepared from the 3-hydroxymethyl-7-phenylacetamido-3-cephem-4-carboxylic acid, the carboxylic acid being protected as a benzhydryl ester and converted to the corresponding 3-formyl compound , These are then allowed to react, according to Scheme 2, with the phosphoranyl compound obtained from the halide compound of formula R CH-X and introduce the desired 7-acylamino group by acyl exchange.

Process E is a further variation of Reaction Scheme 2 using the blocked 7-p-hydroxyphenylglycylamido-3-formyl-3-cephem-4-carboxylic acid as the carbonyl compound.

The 7-phenylacetamidocephdlosporansciure is a suitable starting material because it is readily available. The acetoxy group can be easily enzymatically hydrolyzed using wheat bran as an enzyme source to 7-phenylacetamido-3-hydroxymethyl-ceph-3-em-4-carboxylic acid. The carboxylic acid group may be protected in the form of the benzhydryl ester obtained by treatment of the acid with diphenyldiazomethane. receives. The ester is then treated with phosphorus pentachloride under known reaction conditions resulting in cleavage of the 7-phenylacetyl group and conversion of the 3-hydroxymethyl group to a 3-chloromethyl group. The preparation of the 7-amino-3-chloromethyl-3-cephem-4-carboxylic acid benzhydryl ester according to these methods is illustrated in Examples 1 and 2.

Alternatively, the 7-phenylacl-mandy-j-hydroxymethylceph-3-em-4-carboxylic acid can be converted into the 3-HaJogcMiinet.hy! Compound and '

M / 24 279 -36-

_N 'are then converted into the Phosphoranylverbindung. Subsequently, the reaction is carried out with an aldehyde for the preparation of the substituted 3-vinylcephalosporin according to one of the variants of Reaction Scheme 1.

The cephalosporin-3-carboxaldehyde of the formula QGH = O used as the carboriyl compound in Reaction Scheme 2 is prepared by oxidation of a T-acylamino-S-hydroxymethyl-ceph-3-em-4-carboxylic acid ester as described in U.S. Pat U.S. Patent 3,351,596. Scheme 2 describes the less preferred synthetic route and does not appear to be suitable for preparing the propenyl compounds of formula XIII.

The compounds of formula QCIl = CIlR exist in the cis (Z) and trans (E) configuration. Preferred are the compounds with eis (or Z) -Koniiguration. 'They have greater antibacterial activity than the corresponding compounds with trans (or E) configuration. The compounds of formula XIV are useful as intermediates for the preparation of further cephalosporins of formula QCH = CHR, wherein R represents a methylene group substituted with a nucleophilic residue such as a mercapto, alkylmerodpto, arylmercapto or heteroarylmercapto group, for example, a 1,2,3-triazol-5-yl-mercapto and 2-methyl-6-pyridinylmercaptogruppe. This

OQ method is explained in Example 20. The iodomethyl compounds are the preferred intermediates for the nucleophilic substitution reactions. ,

M / 24 279 -37-

Scheme 1 also includes the preparation of a compound of formula XIV, wherein the corresponding carbonylic ^! Compound of the formula XAIkCHO used instead of the R CHO compound shown.

M / 24 279

example 1

Benzhydryl-3-hydroxymethyl-7β-phenylacetamido-3-cephem- 4-carboxylate (Compound 1) - _l ___ _m

To a suspension of wheat bran (20 g, dry) in phosphate buffer (pH 7, 162.5 ml) is added with stirring at room temperature T-phenylacetamidocephalosporic acid sodium salt (5 g, 12.1 mmol) in one portion. The course of the reaction is followed by HPLC until complete hydrolysis (5 h). The suspension is filtered to remove the wheat bran and the filtrate cooled to 5 to 10 ° C for extractive esterification. To the cooled solution is added methylene chloride (32 ml) and then a 0.5 M solution of diphenyldiazomethane in methylene chloride (24 ml). The pH is then adjusted to 3.0 with 28% phosphoric acid. After 1 h, the temperature of the reaction mixture is allowed to rise to 20 ^° C. Heptane (56 ml) is added slowly and the resulting crystalline title compound is isolated by filtration. The yield is 3.0 g (50%).

B e 1 sp ea I 2

.25. Benzhydryl-7β-amino-3-chloromethyl-3-cephem-4-carboxylate (2)

COOCH (Ph)

35

M / 24 279 -39-

To a slurry of PCl ₁ - (8.3 g; 40 mmol) in CH ₂ Cl ₂ (1000 ml), pyridine (3.2 g; 40 mmol) and the mixture is stirred for 20 min at 20 ⁰ C. At this mixture is added at once under stirring at -40 ⁰ C benzhydryl-3-hydroxymethyl-7-phenylacetamido-3-cephem-4-carboxylate O) (5.1 g; 10 mmol). The mixture is stirred for 15 min at -10 ° C and then allowed to stand for 7 h at -10 to -15 ⁰ C.

To the cooled solution (1-20 ⁰ C) are added propane-1,3-diol (10 ml) and the mixture is allowed to stand for 16 h at -20 ° C and then for 20 min at room temperature with stirring. The resulting solution is washed with ice-water (2 × 20 ml) and saturated aqueous NaCl (10 ml), dried over MgSO 4 and concentrated in vacuo. The gummy residue (12 g) is dissolved in a mixture of CH 2 Cl 2 and n-hexane (2: 1) and chromatographed on a silica gel column (200 g) using the same solvent as the eluent. Those fractions containing the title compound are evaporated in vacuo. The residue is triturated with η-hexane to give the product 2 (2.1 g; 51%); mp. 110 ⁰ C (dec.) Are obtained.

25 IR: ^ ₀₁ , 3400, 2800, 1785, 1725cm " ¹

' ^^ max ** ^{265 1} ^ ( ^E 1c ^{m 16} °) NMR: ^ DMSO-d? + CDCl-

ppm

3.69 (2H, s), 4.43 (2H, s), 5.09 (1H, d, J = 4.5 Hz),

5.24 (1H, d, J = 4.5Hz), 6.87 (1H, s), 7.3 (1OH, m).

Example 3

Benzhydryl-7β- [D-2- (t-butoxycarbonylamino) -2- (p-hydroxyphenyl) acetamido] -3-, chloromethyl-3-cephem-4-carboxylate

(Compound 3) / _

M / 24 279. -40-

i ''. ·

HO

CH ₂ Cl

To a mixture of 20.7 g (0.05 mol) of benzhydryl-7-amino-3-chloromethyl-3-cephem-4-carboxylate (2) and 20 g (0.075 mol) of D-2- (t-butoxycafonylamino) 2- (p-hydroxyphenyl) -acetic acid in 500 ml of dry tetrahydrofuran (THF) are added 15.45 g (0.075 mol) of N, N'-dihydroxyhexyl-1-carbodiimide (DCC) and the mixture is stirred for 2 h at room temperature and then evaporated to the dry. Dissolve the residue in 1 1 of ethyl acetate (AcOEt) and filter off the insoluble dicyclohexylurea. The filtrate is washed with aqueous sodium bicarbonate solution, water and saturated brine, dried over anhydrous sodium sulfate and the solution is evaporated to dryness. The oily residue is on. Chromatography of a silica gel column (Wako gel C-100, 500 g), eluting with 4 1 chloroform and 6 1 1% chloroform-methanol. The desired fractions

, are combined ^ and evaporated to dryness. The oily residue is triturated with ether-isopropyl ether to give 30.6 g (9296) of compound (3).

IR: ^{1 1} cm " ¹ 1790, 1710, 1670, 1500, 1360, 1230, 1150 ΐπείΛ

NMR: 6 ^CdCl 3 ppm 1.45 (9H, s, C-CH ₃ ), 3.4 (2H, br.s, ₂ -H), 4.28 (2H ₁ s, CH ₂ Cl), 4.86 (1H, d, 4.5Hz, 6-H), 5.12 (1H, d, 6Hz, CH-CO), 5.68 (1H, dd, 8 and 4.5 Hz, 7 -H), 6.63 (2H, d, 9Hz, phenyl-H), 6.93 (1H, s, CH-Ph ₂ ), 7.08 (2H, d, 9Hz, phenyl-H), 7.0-7.5 (1 OH, m, phenyl-H).

M / 24 279 -41-

The oily residue can be used without further chromatographic purification in Example "4

- Example 4

Benzhydryl-7β- [D-2- (t-butoxycarbonylamino) -2- (p-hydroxyphenyl) -acetamido] -3-iodoethyl-3-cephem-4-carboxylate (Compound 4)

10. ·.

A mixture of 26.6 g (0.04 mol) of 3_ and 18 g (0.12 mol) of sodium iodide in 400 ml of acetone is stirred for 2 h at room temperature and evaporated to dryness. The residue is extracted with 400 ml of ethyl acetate and the extract is washed with aqueous Na 2 SO 3 PO 4 solution, water and saturated aqueous NaCl solution. After evaporation of the solvent, the residue is triturated with ether-isopropyl ether to give 27 g (89%) of the title compound. If desired, the ethyl acetate solution may be used directly in the next step (compound 5) without isolation of compound 4.

IR: ^^^ cm " ¹ 1790, 1710, 1670, 1500, 1360, 1220, 1150

ITIgIA.

NMR: iS ^CDC1 3 ppm 1.47 (9H, s, C-CH _3), 3.3-3.6 (2H, m, 2H), 4.20 (2H, s, CH _2), 4 , 89 (1H, d, 4.5 Hz, 6-H), 5.12 (1H, d, 6 Hz, CH-CO), 5.68 (1H, dd, 8 and 4.5 Hz, 7 H), 6.62 (2H, d, 9Hz, phenyl-H), 6.92 (111, s, CHPh ₂ ), 7.08 (2H, d, 9Hz, phenyl-H),

30 7-7.5 (1 OH, m, phenyl-H).

Example 5 '

Benzhydryl-7β- [D-2- (t-butoxycarbonylamino) -2- (p -hydroxyphenyl) -acetamido] -3- (triphenylphosphonio) -methyl-3-cephem -4-carboxylate-iodide (compound 5)

M / 24 279

CHCONH I NH

CO ₂ C (CH ₃ )

CU ₂ CH

A mixture of 15.1 g (0.02 mol) of 4 and 15.7 g (0.06 mol) of triphenyl phosphine in 200 ml of ethyl acetate is stirred for 1 h at room temperature. The resulting precipitate is filtered off to give 17.4 g (85.5%) of 5, with a melting point of 170 to 180 ° C. Concentrate the filtrate to 100 ml and dilute the concentrate with 500 ml of ether to give a second fraction (1.1 g). The total yield is 18.5 g (91%). The overall yield of 5, starting from compound 2, is 74.5%. This yield can be increased to 87.5% by omitting the purification and isolation steps as indicated above.

I ^R '· ^^ cm * " ¹ 1780, 1670, 1490, 1420, 1350, 1240, 1150, 1090

NMR: 6 ^DMS0 ppm 1.42 (9H, s, C-CH _3), 3.45 (2H, br.s, 2H), 5 to 5.4 (3H, m, 3H and 6- H), 5.7 (1H, m, 7-H), 6.63 (2H, d, 9Hz, phenyl-H), 7.1-7.45 (12H, m, phenyl-H), 7, 5-7.9 (15H, m, phenyl-H)

Elemental analysis: for Cc ₂ H ^ g

Calculated: C 61.36% H 4.85% Found: 61.26 4.82

N 4.13% 4.11

S 3.15% 3.92

M / 24 279

10

Example 6

Benzhydryl 7ß- [D-2- (t-butoxycarbonylamino) -2- (p-hydroxyphenyl) acetamido] -3 - [(2) -1-propen-1-yl] ceph-3-em-4- carboxylate (compound 6)

C lt = CHCH-

CO ₂ C ₁ (CH ₃ )

15 20 25 30 35

To a solution of 1. 8 g (1.77 mmol) in 100 ml of chloroform is added 100 ml of water containing 2 ml (2 mmol) of sodium hydroxide and the mixture is shaken for 5 min. The organic layer is separated, washed with water and dried over anhydrous sodium sulfate. The chloroform solution is filtered and the filtrate is concentrated under reduced pressure to 50 ml. To the concentrate is added 1 g of acetaldehyde and the mixture is stirred for 2 h at room temperature and then evaporated to dryness. The oily residue is chromatographed on a silica gel column (Wako-gel C-200, 50 g) eluting with chloroform and chloroform-methanol (99: 1). The "desired fractions are combined and the solvent evaporated to give 318 mg (28%) of compound 6, mp. 120 to 130 ⁰ C (dec.).

IR: v> 55 cm " ¹ 1780, 1670, 1710, 1490, 1360, 1210, 1150

NMR: S ^CDCl 3 pp _m 1.3-1.5 (12H, m, C-CH ₃ ), 3.22 (2H, br.s, 2-H), 4.90 (1H, d, 4, 5Hz, 6-H), 5.15 (1H, br.d, CH-CO), 5.5-6.1 (3H _1m , CH = CH and 7-H), 6.63 (2Hy d , 9Hz, phenyl-H), 6.91 (1H, s, CH-Ph), 7.09 (2H, d, 9Hz, phenyl-H), 7.2-7.5 (1OH, m, phenyl-H).

M / 24 279

Example 7

Sodium 7β- [D-2-amino-2- (p -hydroxyphenyl) acetamido] -3 - [(Z) -1-propen-1-yl] -3-cephem-4-oxyloxylate (Compound 7 »BMY -28100 sodium salt)

CH = CHCH.

(ice)

COOH

A mixture of 318 mg (0.48 mmol) of 6 and 2.5 ml of trifluoroacetic acid (TFA) is stirred for 1 h at room temperature and then diluted with 50 ml of ether and 50 ml of isopropyl ether. The deposited precipitate is filtered off and washed with ether to give 188 mg (77%) of the trifluoroacetate of 7, which is dissolved in 2 ml of methanol (MeOH). To this solution is added 2 ml (2 mmol) of a solution of sodium 2-ethylhexanoate (SEH) in ethyl acetate, and the mixture is diluted with 30 ml of ethyl acetate to precipitate the precipitate. The precipitate is filtered off, washed with ether and in vacuo over Ρ? ^Ο 5, whereby 144 mg (73%, based on compound 6) of crude compound 7 is obtained. The crude product (135 mg) is dissolved in 10 ml of water and the solution is chromatographed on a column (25 mm × 100 mm) using about 20 ml PrepPak-500 / Cg (Waters) as filler. The column is eluted with water, the eluate containing the desired product is concentrated to 5 ml and lyophilized to give 93 mg (69%) of compound 7, mp 200 ° C (gradual decomp.). The estimated purity is 60%

35 (by HPLC).

M / 24 279 -45-

i O JPÜ ^{cra "1} 1760, 1660, 1590, 1400, 1360, 1250

Slide X

UV: / \ Phosphatpuff er pH 7 _{M1 ( b) 22?} ( ₁₁₃₀ 0), 280 (8200)

i ^D 2 ° ppm 1.65 (3H, d, 6Hz, -C-CH ₃ ), 3.21 (1H, d,

18 Hz, 2-H), 3.52 (1H, d, 18 Hz, 2-H), 5.12 <1H, · d, 4.5 Hz, 6-H), 5.68 (1H, d, 4.5 Hz, 7-H),

5.5-5.9 (1H, m, vinyl-H), 5.95 (1H, d, 11.5Hz, vinyl-H), 6.94 (2H, d, 8Hz, phenyl-H) , 7.36 (2H, d, 8H7, phenyl-H).

Example 8,. .

7β- [D-2-amino-2- (p -hydroxyphenyl) -acetamido] -3 - [(E) -1-propen-1-yl] -3-cephera-4-carboic acid (Compound 8,

BB-S1067) ; .

11.9 g of the crude product 7 prepared according to Example 7 are dissolved in 50 ml of 0.01 M phosphate buffer (pH 7.2) -methanol (85:15) in the state in which they are before the chromatographic purification adjust the pH of the solution to 6 with 6N hydrochloric acid.

This solution is by means of high pressure liquid chromatography (HPLC) (PrepPak-500 / _C18, System 500, V / Ater) using a 0.01 M phosphate buffer (pH 7.2) containing methanol 159O treated. The eluate is analyzed by analytical HPLC to find that the first 4 1 fraction contains the cis isomer BMY-28100. The second 1 1 fraction contains the transisomer and is concentrated to 500 ml. The concentrate is adjusted to pH 3 by means of dilute hydrochloric acid and chromatographed on an HP-20 column (100 ml) and eluted with 1 l each of water and 30% strength

M / 24 279 -46- '

Methanol. The latter eluate having a volume of about 300 ml is concentrated to 10 ml and lyophilized to obtain 290 mg of the crude trans isomer (5596 purity). This material is dissolved in 100 ml of 50% methanol and treated with charcoal. The FII entered is concentrated to a volume of 20 ml and allowed to stand overnight at 5 ⁰ C. The product crystallizes in the form of colorless prisms, which are filtered off and dried in vacuo. This gives 129 mg, mp. 23O ⁰ C (dec.).

IR: v> J ^ cm ~ ¹ 1760, 1680, 1590, 1550, 1520, 1450, 1390, 1350, 1240

⁷⁾ mn. (t) 228 (13000), 292 (16900)

NMR: o ^D 2 O ^{+ Na} 2 ^C0 3 ppm 1.89 (3H, d, 6 Hz, C = C-CH ₃ ),

3.60 (2H, s, 2-H), 5> 13 (1H, d, 4.5Hz, 6-H), 5.20 (1H, s, CH-CO), 5.68 (1H, d, 4.5 Hz, 7-H), 5.99 (1H, dq, 16 and 6 Hz), 6.54 (1H, .d / 16 Hz), 6.98 (2H, d, 9 Hz, Phenyl-H), 7.41 (2H, d, 9 Hz,

25-phenyl-H).

Example 9

Crystalline 7β- [D-2-amino- (p-hydroxyphenyl) -acetamido] -3 - [(Z) -1-propen-1-yl] -3-cephem-4-carboxylic acid ( Compound 9 "BMY-28100 ) '

The first 4 l fraction obtained according to the preparative HPLC method described in Example 8 and containing the cis isomer (BMY-28100) is concentrated to a volume of 2 l. The pH of the concentrate provides

M / 24 279 -47-

one with dilute hydrochloric acid to 3. The solution is added to a column containing HP-20 (11) and the column is washed with 6 l of water until the pH of the effluent is 7. The column is then eluted with 4 L of 30% aqueous methanol. The eluate is analyzed by HPLC. The appropriate fractions are combined (about 2.5 l) and concentrated to 50 ml at a temperature of less than 40 ^° C. and under reduced pressure. It forms a crystalline precipitate. The concentrate is cooled for 2 h to ^{0 0} C and filtered from the crystalline precipitate, washed with square aqueous acetone, then with 100% acetone and then dried in vacuo over P ₂ ° 5 » ^with 4.09 g of pure , crystalline, desired compound, m.p. 218 to 220 ⁰ C (dec.) Obtained in the form of colorless prisms, which have a purity of 95% according to HPLC analysis. ,

20 '

^IR: ^ ί ^ χ ^cm " ¹ 1750, 1680, 1560, 1520, 1460, 1390, 1350, 1270, 1235

^{UV: Λ} ^ ^OS P ^{has buffer (pH 7)} nm ( i ) 228 (1.2300), 279 (9800)

IiMR: t5 ^D 2 ^{O + NaHCO} 3 ppm 1.71 (3H, d, ^6Hz , C-CH ₃ ), 3.27 (1H, d, ^{18Hz, 2H} ), 3.59 (1H, d , 18 Hz, 2-H) /

5.18 (1H, d, 4.5Hz, 6H), 5.22 (1H, s, CHCO),

5.73 (1H, d, 4.5Hz, -7H), 5.5-6.0 (1H, m, CH = C),

6.02 (1H, d, 11 'Hz, CH = C), 6.98 (2H, d, 9 Hz,

Phenyl-H), 7.41 (2H, d, 9Hz, phenyl-H) '. Elemental analysis: for C ₁₈ H ₁ ON ₃ OcS-I ^ H ₂ O, calculated: C 54.26% H 5.06% N 10.55% S 8.05% Found: 54.15 5.13 10.30 8.38

54.19 5.08 '10.42 8.04

M / 24 279 »48-

The mother liquor obtained after the crystallization is concentrated to a volume of 10 ml and treated with 20 ml of acetone. When the solution is stored overnight in the refrigerator, a crystalline precipitate forms, which is filtered off and dried in vacuo over PpOc; the amount is 670 mg (purity 90% according to HPLC). 560 mg of this material are dissolved in 200 ml of 50% aqueous methanol and the solution is treated with 0.5 g of activated charcoal and filtered off. The filtrate is concen- at reduced pressure and at 40 ⁰ C to a volume of 20 ml - trated and then held for 5 hours at 5 ° C. The product crystallizes, it is filtered off, washed with acetone and dried in vacuo over Pp ^ 5, to give 227 rag of crystalline BMY-28100 (98% purity, determined by HPLC). Lyophilization of the mother liquor gives 181 mg BMY-28100, which has a purity of 95% (HPLC). ,

Example 10

Diphenylmethyl-7β- [D-2- (t-butoxycarbonylamino) -2- (phydroxyphenyl) acetamido] -3-vinyl-3-cephem-4-carboxylate (Compound 10) -

A solution of 3 g (2.95 mmol) of benzhydryl 7- [2- (Nt -butoxycarbonylamino) -2- (p-hydroxyphenyl) acetamido] -3- (triphenylphosphine) -methyl-3-cephem- 4-Carboxylate iodide (5) in 50 ml of chloroform is shaken for 1 minute with a mixture of 3 ml (3 mmol) of 1 N NaOH and 50 ml of water at room temperature. The organic layer is separated after addition of saturated NaCl-r solution (20 ml) and washed with water (3 x 30 ml). To the organic solution is added with vigorous stirring and under cooling with water 2.5 ml of 39% aqueous formaldehyde and stirred

M / 24 279 -49-

another 20 min. The organic layer is separated, dried over anhydrous Na ₂ SO ₄ and concentrated in vacuo. The concentrate is placed on a silica gel column which is eluted with CHCl ₃ (600 ml) and 2% MeOH in CHCl ₃ (800 ml) to give 850 mg (45%) of the title compound. TLC: Rf 0.48 [silica gel, MeOH-CHCl ₃ (1:10)]

 10

At game 11

7β- [D-2-Amino-2- (p -hydroxyphenyl) -acetaniido] -3-vinyl] -3 -cephem-4-carboxylic acid (Compound 11, BB-S1064)

A mixture of 850 mg (1.32 mmol) of compound 10 and 5 ml of 90% aqueous trifluoroacetic acid (TFA) is allowed to stand for 1 h at room temperature and concentrated to about 1 ml in vacuo. The concentrate is triturated with 20 ml of diisopropyl ether to give 679 mg of a yellow powder, which is dissolved in 3 ml of methanol and then diluted with 30 ml of water. The solution is passed through an HP-20 column (50 ml), which is washed with 200 ml of water and eluted with 250 ml of 30% methanol. The eluate containing the desired compound was concentrated and lyophilized to yield (31%) is obtained • 197 mg of the title compound with an estimated purity by HPLC of 60% and having a melting point of 190 ⁰ C (dec.). , * '

30 IR: *) ίί ί cm ~ ¹ 1760, I68O, 1615-1570, 1520

_m . _Λ phosphate buffer (pH 7) _{nm (f) 2} 28 (135OO), 283 (14400)

NMR: cf ^D 2 ° ppm 3.6 (2H, s, SCH ₂ ), 5.51 (1H, d, 5Hz, 6-H), 5.73 (1H, d, 5Hz, 7-H), 7.03 (2H, d, 8Hz, phenyl -H), 7.45 (2H, d, 9Hz, phenyl-H).

(. "

M / 24 279 -50-

ι. .- '".'.

Example 12

Diphenylmethyl-7β- [D-2- (t-butoxycarbonylamino) -2- (phydroxyphenyl) acetamido] -3 - [(Z) -1-buten-1-yl] -3-ceph- em-4-carboxylate ( Connection: 12)

A solution of 3 g (2.95 mmol) of compound 5 in 50 mL of CH 2 Cl 2 is mixed with a mixture of 3 × 2 mL (3.2 mmol) 1N NaOH and 50 mL of water and the mixture is shaken at room temperature for 3 minutes. The organic layer is separated, washed with water (3 x 30 ml) and saturated NaCl solution and dried over anhydrous NapSOr. To this solution is added 1.71 g (29.5 mmol) of propionaldehyde. The mixture is stirred at room temperature overnight and concentrated at reduced pressure. ^r The concentrate is placed on a silica gel column eluted with 1 to 2% methanol in CHCl ₃ . Those fractions which show a spot at Rf 0.30 (TLC, MeOH-CHCl, = 1:10) ^ are combined and evaporated to give 1.08 g (55%) of the title compound.

IR '· J ¹ OSi cm " ¹ 1780, 1680, 1500. 25

Example 13

Sodium 7ß- [D ~ 2 ~ amino-2- (p-hydroxyphenyl) acetamido] -3 - [(Z) -1-buten-1-yl] -3-cephem-4-carboxylate (Compound 13, BB -S1058 sodium salt). .

30,

A solution of 1.08 g (1.61 mmol) of Compound 12 in 11 ml of TFA containing 1% water is allowed to stand at room temperature for 1 h. The mixture is concentrated in vacuo to about 2 ml and the resulting syrup is triturated with about 20 ml of diisopropyl ether to give 796 mg of a yellow

M / 24 279 -51-

Powder receives. The powder is dissolved in 3 ml of methanol and the solution is treated with 3 ml of 0.8 M SEH in ethyl acetate (AcOEt) to form a precipitate, which is filtered off, washed with diisopropyl ether and dissolved in 5 ml of water. The solution is placed on a column filled with prepPAK-500 / C _i8 (Waters). The column is then washed with water and alumina sequentially with 10% methanol, 20% methanol and 30% methanol. The desired fractions (monitored by HPLC) are combined, concentrated and lyophilized to give 118 mg (9.4%) of the ti te compound with an estimated purity of 55% (by HPLC). This compound turns dark when heated to> 180 ° C in a glass capillary.

^IR: ^ £ £ ^cm ~ ¹ 1755, 1660, 1580

^{m: X} ^ ° ^{Sphat buffer (pH 7)} nm it ) 228 - (10900), 278 (7200)

'NMR: c $ ^D 2 ° ppm 0.81 (3H, t, 7.5 Hz), 1.7-2.2 (2H, m), 3.25 (2H, ABq), 5.01 (1H , d, 5Hz), 5.50 (1H, d- t, 7.5 and 12Hz), 5.58 (1H, d, 5Hz), 5.78 <1H,

d, 12 Hz), 6.86 (2H, d, δ Hz), 7.26 (2H, d, 8 Hz) ".

Beis-p'iel 14

Diphenylmethyl-7β- [D-2- (t-butoxycarbonylamino) -2- (phydroxyphenyl) acetamido] -3 - [(Z) -3-phenyl-1-propen-1- yl] -3-cephem-4 carboxylate (compound 14)

A solution of 3 g (2.95 mmol) of Compound 5 in 50 ml of CH 2 Cl 2 is shaken for 1 minute with a mixture of 3.2 ml (3.2 mmol) of 1 N NaOH and 50 ml of water. The organic

M / 24 279 -52-

Layer is separated after addition of a saturated NaCl solution (20 ml), washed with water (3 x 30 ml) and saturated NaCl solution and dried over anhydrous NapSO. dried. Then 7.2 g (30 mmol) of 5 Obigen phenylacetaldehyde are added to this solution and the mixture is stirred overnight at room temperature. The reaction mixture is then concentrated in vacuo and the concentrate is purified on a silica gel column (75 g) using 1% MeOH / CH 2 Cl 2 to give 800 mg (37%) of the title compound. Thin Layer Chromatography (TLC): Rf 0.33 (silica gel, MeOH-CH 2 Cl 2, 1:10); IR (KBr): 1780, 1710-1680 cm "\

This compound is used without further purification in Example 15.

•, Example 15

7β- [D-2-Amino-2- (p -hydroxyphenyl) -acetamido] -3 - [(Z) -3-phenyl-i-propen-i-yl] i-cephem-1-carboxylic acid (Compound 15, BB-S1O76). ____ ".

A solution of 800 mg (1.09 mmol) of Compound 14 in 4 ml of 90% TFA is allowed to stand for 2 h. Concentrate the reaction mixture and triturate the concentrate with diisopropyl ether to give 490 mg of a yellow powder. A solution of the powder in 2 ml of methanol is mixed with 20 ml of water and the mixture is poured onto an HP-20 column (50 ml), which is washed with water (250 ml) and washed successively with 30% methanol (250 ml). and 75% methanol (300 ml) is eluted. The eluate with 75% methanol is concentrated and lyophilized to give 302 mg of the crude product, which is dissolved in 10 ml of 75% methanol and applied to a column using PrepPAK-500 / C (Waters) (80 ml) filling

M / 24 279 -53-

is chromatographed. Elution of the column with 75% methanol gives 158 mg (315 °) of the desired product. The estimated purity (by HPLC) is 65 °. The product turns dark when heated in a capillary tube> 175 ° C.

^IR : v? 1 cm. " ¹ 1760, 1680., 1600-1580, 1520 ^m: λ ^ ^{sphat buffer (pH 7)} nm (£) 280 (8900)

(5/1)

_{ppm 4> 45 (2H, d> 4} ^

4.87 (1H, s, CHND ₂ ), 6.7 (2H, d, 9Hz, Ph), 6.9-7.5 (7H, ra, Ph).

Example 16 .

Diphenylmethyl-7β- [D-2- (t-butoxycarbonylamino) -2- (phydroxyphenyl) -acetamido] -3 - [(Z) -3-methoxy-1-propen-1- yl] -3-cephem-4 carboxylate (compound 16)

A solution of 3.0 g (2.95 mmol) of compound 5 in CH 2 Cl 2 (100 ml) is treated with a mixture of 2N NaOH (1.8 ml) and water (100 ml) at room temperature for 5 min. The organic phase is separated, washed with water (50 ml) and aqueous NaCl solution (50 ml), dried and concentrated to about 10 ml. The resulting red ylide solution is treated with methoxyacetaldehyde (1.3 ml, 15 mmol) for 15 min at room temperature. After stripping off the solvent, the residue is chromatographed on a silica gel column (100 g) using toluene-AcOEt (3: 1 and 1: 1) as the eluent to give the title compound (750 mg, 38%).

M / 24 279 -54-

NMR: J ^CDC1 3 ^{+ D} 2 ° ppm 1.45 (9.H, s, t-Bu), 3.15 (3H, ^s,:

, OCH ₃ ), 3,27 (2H, s, 2-CH ₂ ), ca. 3,5 (2H, m, -CH ₂ -OMe), 4,90 (1H, d, 5,0 H7, 6 H), 5.12 (1H,

s, -CH-ND-), ca. 5.5 (1H, m, = CH-CH ₂ -), 5.72 (1H, d, 7-H), 6.18 (1H, d, 12 Hz , -CH = CH-CH ₂ -), 6.65 and 7.10 (each 2H, each d> HO-Ph-), 6.90 (1H, S, -CHPh ₂ ), 7.3 (10H, s, Ph). 10

Example 17

7β- [D-2-Amino-2- (p -hydroxy-phenyl) -acetamido] -3- [(Z) -3-methoxy-1-propen-1-yl] -3-cephem-4-carboxylic acid, Compound 17. BB -S1O92)

Compound 16 is deblocked by treatment with TFA (3 ml) for 1 h at room temperature. Evaporation of the solvent followed by precipitation from isopropyl ether gives the trifluoroacetate, which is purified by HP-20 column chromatography. The column is washed with H ₂ O (500 ml) and eluted with 30% MeOH (500 ml) to give 350 mg (75%) of the desired product with an estimated purity of 90% (by HPLC), mp 160 ⁰ C (Zers.), Receives.

''

IR: v> JP ^ cm " ¹ 3400, 3180, 1760, 168O

_{: Λ} phosphate buffer (pH 7) ^^ ₂ 28 (115OO), 279 (9400)

NMRtif ^ ⁰ ppm 3.40 (3H, s, OCH _3), 3.40 (2H, ABq, 2-CH _2). 4.0 (2H, m, -CH ₂ OMe), 5.19 (1H, d, 4.5Hz, 6-H), 5.25 (1H, s, -CH-IO ₂ ), 5.77 (1H, d, 7-H), ca. 5,8 (1H, m, ^ CH-CH ₂ -), 6,20 (1H, d, 11 Hz ₁ -CH = CH-CH ₂ ), 7, 05 and 7.45 (each 2H, each d, HO-Ph-).

.

M / 24 279 -55-

1 .

Example 18

Diphenylmethyl-7βr [D-2- (t-butoxycarbonylamino) -2- (phydroxyphenyl) -acetamido] -3 - [(Z) -3-chloro-1-propen-1-yl] -3-cephem-4-carboxylate (Compound 18)

A solution of compound 5 (5.0 g, 4.9 mmol) in CH 2 Cl 2 (100 mL) is stirred for 5 min at room temperature with a mixture of 2N NaOH (2.9 mL, 5.8 mmol) and water (100 mL ). The organic phase is separated and washed with water (50 ml) and saturated NaCl solution (50 ml) and dried over anhydrous Na ₂ SO 4. The filtrate is concentrated to about 20 ml and treated with chloroacetaldehyde (2.0 ml, 25 mmol). The mixture is stirred for 30 min at room temperature and evaporated in vacuo. The syrupy residue is chromatographed on a silica gel column (100 g) using toluene-AcOEt (3/1) as the eluant, using the

20 title compound 18 is obtained (900 mg, 27%).

NMR. f, ^CDC1 3 ^{+ D} 2 ° ppm 1.45 (9H, s, t-Bu), ca. 3.3 (2H, m, 2-CH ₂ ), 3.5-4.0 (2H ₁ m, -CH ₂ -Cl), 4.92 (1H, d, 5.0Hz, 6-H), 5.12 (1H, s, -CH-ND-), ca. 5.7 (2H, m, 7-H and = CH-CH ₂ ), 6.15 (1H, d, 11 Hz, 3-CH = CH-CH ₂ -), 6.63 u.7.10 (2H each, each d, HO -Ph-), 6.89 (1H, s, CHPh ₂ ), 7.3 (1OH, s, Ph).

Deblocking this compound with TFA according to the methods described in the previous examples (e.g., Examples 7, 11, etc.) gives the 7- [D-2-. Amino-2- (p-hydroxyphenyl) acetamido] -3 - [(Z) -3-chloro-1- propen-1-yl) -3-cephem-4-carboxylic acid.

M / 24 279 -56-

B ile le I 19

Diphenylmethyl-7β ~ [D-2- (t-butoxycarbonylamino) -2- (phydroxyphenyl) -acetamido] -3 - [(E) -3-iodo-1-propen-1-yl] -3-cephem-4 carboxylate (compound 19)

A mixture of compound 18 (900 mg, 1.3 mmol) and NaI (590 mg, 3.9 mmol) in acetone (18 ml) is stirred for 1 h at room temp era. After evaporation of the solvent, the residue is dissolved in AcOEt (100 ml), washed successively with water, aqueous Na ₂ S ₂ O ^ - and aqueous NaCl solution, the solution is dried and the solvent evaporated to give the title compound (1.02 g) receives.

NMR: ir ^CDei 3 "" ^{: D} 2 ⁰ ppm 1.45 (9H, s, t-Bu), ca. 3.4 (2H, m, 2-CH ₂ ), ca. 3.8 (2H, m , -CH ₂ -J), 4.90 (1H, d, 5.0He, "6-H), 5.14 (1H, s, -CH-ND-), 5.73 (iH _t d, .7-H), ca.5,5-6,0 (1H, m, = CH-CH ₂ -), 6.68 and 7.10 (each 2H, each d, HO-Ph-). 6 , 78 (1H ^ d, 15 Hz, 3-CH = CH-CH ₂ -), 6.99 (1H, s, CHPh ₂ ), 7.35 (1H, s, Ph).

25 B el s ρ ie.1 20

Diphenylmethyl-7β-> [D-2- (t-butoxycarbonylamino) -2- (phydroxyphenyl) acetamido] -3- [3 - (iH-1,2,3-triazol-5-yl) thio] 1-propen-1-yl] -3-cephem-4-carboxylate (Compound 20)

To a solution of compound 19 (1.0 g, 1.3 mmol) in ethyl acetate (20 mL) is added propylene oxide (0.27 mL, 3.8 mmol) and 0.1 M (1H-1,2,3 Triazol-4-yl) thiol in ethyl acetate (19 ml). The mixture is stirred for 30 min at room temperature and evaporated under reduced pressure. The syrupy residue is chromatographed on a silica column C-200 (50 g). The wished

M / 24 279 -57-

Product is eluted with CHCl ₅ -MeOH (10: 1) to give 800 mg (83%) of the title compound. "

' mRiS ^CDC1 3 ^{+ H} 2 ° ppm 1.45 (9H, s, t-Bu), ca. 3.3 (4H, m, 2-CH ₂ -, and -CH ₂ -S), 4.80 ( 1H, d, 5.0Hz, 6-H), 5.20 (1H, s, -CH-ND-), 5.70 (1H, d, 7-H), ca. 5.95 (1H, m, = CH-CH ₂ -), 6.68 (2H, d, HO-Ph-), 6.90 (1H, s, -CHPh ₂ ), 7.25 (1OH, s, Ph), 7, 52 (1H, s, triazole-4-H).

Example 21 7β- [D-2-Amino-2- (p -hydroxyphenyl) acetamido] -3- [3- (1H-

Bonic acid (compound 21. BB-S1091)

A mixture of compound 20 (800 mg) and TFA (2 ml) is kept at room temperature for 1 h and then evaporated to dryness. Isopropyl ether is added to the residue to give a yellow precipitate (600 r.p.), which is dissolved in water (1 ml) and applied to an HP-20 column (100 ml). The column is washed with water (500 ml) and eluted with 30% MeOH followed by 50% MeOH. The fractions containing the desired compound are combined, evaporated and lyophilized to give 170 mg (33%) of the desired product with an estimated purity of 50% (by HPLC), mp. 180 ⁰ C (dec.), It -

30 stops.

IR "- Sv cm" ¹ 3360, 3280, 1755, 1670

: λ P ^ ^os P ^has P ^buffer nm (£) 235 (14100), 252 (12300)

ΠΙ Cl Λ

M / 24 279 -58-

NMR: δ ^D 2 ^{0 + DC 1} ppm ca. 3.4 (4H, m, 2-CH ₂ T, .- CH ₂ -S-), 5.43 (1H, d, 4.5 Ηζ, -6 -Η), 5.15 (1H, s, -CH-ND ₂ ), ca. 6.0 (211, m, 7-H and -CH-CH ₂ -), 6.70 u. .7,15

(each 2H, each d "HO-Ph"), 8.05 (1H, s,., triazole-4-H). ,

Example 22

Behzhydryl-7β- [D-2- (6-butoxycarbonylamino) -2-phenylacetamido] -3- (triphenylphosphonio) -methyl ^ -cephera -carboxylate-iodide (Compound 22) . _____

A mixture of 14.5 g (0.0196 mmol) of benzhydryl-7- [D - (-) - a- (t-butoxycarbonylamino) -a-phenylacetamido] -3-iodo-methyl-3-cephem-4-carboxylate and 5.24 g (0.02 mol) of triphenylphosphine in 300 ml of ethyl acetate is stirred for 2 hours at room temperature. 200 ml of ether are added to the reaction mixture, forming a precipitate which is filtered off and washed with ether. The title compound is concentrated to 1, 3 g (739), the filtrate is concentrated to 50 ml and the concentrate is diluted with ether to give 2.4 g of a second fraction of the product. 7g (85%) 25

'IR:> ¹ ^ cm " ¹ 1780, I69O, 1480, 1420, 1350, 1240, 1150. max

Example 23

Benzhydryl 7ß- [D-2- (6-butoxycarbonylaraino) -2-phenylacetamido] -3 - [(Z) -I-propene-1-YLJ-3-cephem-4-carboxylate

(Connection 23) - »« _ »» _-- "« »-_-__-> -.

To a solution of 5 g (5 mmol) of compound 22 in 200 ml of chloroform, a mixture of 100 ml of water and 5 ml (5 mmol) of sodium hydroxide is added and the mixture is shaken.

M / 24 279 -59-

3 min. The organic layer is separated, washed with water and saturated NaCl solution and dried over anhydrous magnesium sulfate. The chloroform solution is filtered, the filtrate is concentrated under reduced pressure to 100 ml. 3 ml of acetaldehyde are added to the concentrate and the mixture is stirred at room temperature for 1.5 h and evaporated to dryness. The oily residue is chromatographed on a silica gel column (Kieselgel 60, 50.g) eluting with chloroform. The desired fractions are combined and concentrated to dryness. The residue is triturated with n * hexane to give 990 mg (31%) of the title compound (23).

15 receives.

:: v> £££ cm ^"1 in 1780, 1710, 1660, 1510, 1490, 1360, 1240, 1210, 1150

NMR: ^ ^CDC1 3 ppm 1.3-1.5 (12H, m, -C-CH _3), 3.22 (2H, s, 2H), 4.93 (1H, d, 4.5 H7 , SU) ₉ '5.23 (1H, d, 8Hz, CH-CO), 5.5-6.2 (3H, m, 7-H and vinyl-H), 6.94 (1H, s , CHPh), 7.2-7.5 (15H, m, phenyl-K).

25 Example 24

Sodium 7β- [D-2-amino-2-phenylacetamido] -3 - [(Z) -I- propenyl] -3-cephem-4-carboxylate (Compound 24, BB-S1O65)

A mixture of 0.94 g (1.47 mmol) of compound 23 and 3 ml of TFA is stirred for 30 minutes at room temperature and then the mixture is diluted with 50 ml of a 1: 1 mixture of ether-isopropyl ether, giving ca.800 Precipitate mg of a precipitate, which is filtered off and dissolved in _v 3 ml of methanol. To the solution is added 4.5 ml (4.5 mmol) of 1 M sodium 2-ethylhexanoate (SEH) in ethyl acetate, and the mixture is diluted successively with 50 ml

M / 24 279 -60 «

Ether and 50 ml of isopropyl ether. Filtration of the precipitate gives 710 mg of crude product 24, which is dissolved in 20 ml of water and chromatographed on a column with 50 ml PrepPAK / C * ₈ (Waters). The column is eluted with water and 10% methanol. The fractions containing the desired product are collected following the course of the chromatography by HPLC, concentrated to 5 ml and lyophilized. Man he ¹ -

holds 182 mg (31%) of the desired product, Fp.200 ⁰ C. with a purity estimated by HPLC of 50%.

^IR : ^ ¹ SZ cm " ¹ 1760, 1660, 1600, 1400, 1180, 1100

Uj ei / C

( ^{pH 7} nm (£) 282 (5500)

NMR: d ' ^D 2 ° ppm 1.60 (3H, d, 6Hz, -C-CH ₃ ), 3.12 (1H, d,

18 Hz, 2-H), 3.48 (IH, d, 18 Hz, 2-H), 5.03 (IH, d, 4.5 Hz, 6-H), 5.62 (1H, d, 4.5Hz, 7-H), 5.93 (1H, d, 10Hz, vinyl-H), 5.2-5.8 (1H, m, vinyl-H), 7.41 (5H, s , Phenyl-H).

Example 25

5 Benzhydryl-7β- [-D-2- (6-butoxycarbonylamino) -2-phenylacetamido] -3 - [(Z) -3-chloro-1-propen-1-yl] -3-cephem-4- carboxylate (Compound 25)

To a solution of 2 g (2 mmol) of compound 22 in 50 ml of chloroform is added 50 ml of water containing 2 ml (2 mmol) of sodium hydroxide and the mixture is shaken for 3 min. The organic layer is separated and washed successively with water and saturated NaCl solution. The dried chloroform solution is concentrated under reduced pressure to 30 ml. To the concentrate

M / 24 279 -61-

Add 2 ml of chloroacetaldehyde and stir the mixture for 1 h at room temperature. It is washed with water and then with saturated NaCl solution. The organic solution is dried and evaporated to dryness. The resulting oily residue is chromatographed on a silica gel column (Wako-gel C-200, 50 g) eluting with chloroform. The desired fractions are collected and evaporated to dryness to give 534 mg of crude product.

IR: v> cm " ¹ 1780, 1710, 1660, 1500, 1490, 1360,

1240, 1210, 1150. 15

The structure of this sample is not confirmed due to the poor NMR spectrum.

Example 26

Sodium 7β- (D-2-amino-2-phenylacetamido) -3 - [(Z) -3-chloro-1-propen-1-yl] -3-cephem-4-carboxylate (Compound 26, BB-S1066 ) _{; t} ; __

j A mixture of 472 mg (0.7 mmol) of compound 25 and 1.5 ml of TFA is stirred for 15 min at 10 to 15 ⁰ C and diluted with 30 ml of a mixture of ether and isopropyl ether (1: 1) to give 330 mg of a pale yellow precipitate, which is filtered off. To a solution of the precipitate in 3 ml of methanol is added 2 ml (2 mmol) of SEH in ethyl acetate and the mixture is diluted with 50 ml of ethyl acetate. The resulting precipitate is filtered off and washed with ether to give 244 mg of crude product. A solution of the crude product in 10 ml of water is chromatographed on a column using 50 ml of a PrCpPAK-SOOZc _1ö (Waters) filling.

M / 24 279

The column is eluted with water and 10% methanol. The desired fractions of IG ^ methanol are combined, concentrated to 5 ml and lyophilized to 200 ⁰ C (allmähl. ZerSe) receives 60 mg of a solid product, mp ". .

cm ™ ¹ 1760 _f 1660, 1630, 1360, 1120, 1070

- . _K phosphate buffer _{m (&) 2} 43 (1270Ö), 200Sch. (4200).

IIIclX

7β- [D (-) - 2-Amino-2-phenylacetainido] -3 - [(Z) -I-propene-1-yl] -3-cephem-4-carboxylic acid (Compound 24, BB-S1O65 'in zwitterionic shape

CH = CHCH

(Z)

Diphenylmethyl-7β- [D-2- (t-butoxycarbonylamino) -2-phenyl ™ acetamido] -3- (1-propenyl) -3-cephera-4-carboxylate (Compound 23; 1.5 g; 2: _9: 34 mmol ) is treated with 3 ml of trifluoroacetic acid and the mixture is stirred for 20 minutes at room temperature. It is diluted with 100 ml of ether to obtain 1 »15 g (96%) of the crude trifluoroacetate of BB-S1O65.

IR: \? ^ Z cm " ¹ 1760, 1670, 1200, 1130

^. _λ phosphate buffer (pH 7) ^ _{u) 283} 35

M / 24 279 -63-

The trifluoroacetate (1.1 g, 2.25 mmol) is dissolved in 20 mL of water and the solution is chromatographed on a column using 100 mL of PrOpPAKZC ₁₈ (Waters). The column is eluted with water, 10% methanol and 30% methanol. The eluates with $ 30 methanol are concentrated to 10 ml. The crystalline product precipitates, it is isolated and washed with acetone and dried in vacuo over PpOc. This gives 505 mg (4690) of pure BB-S1O65 (in zwitterionic form), mp. 180 to 183 ⁰ C (dec.), With an estimated purity of 95%.

IR: J .J *! cm " ¹ 1750, 1690, 159Q, 1400, 1350 UV: λ phosphate buffer (pH 7) ^ _{0 282 (8Q00)}

IücLX.

IücLX.

NMR: ^ D ₂ ⁰⁺ NaHGO _{3 ppffl 1} ^ ₅₈ ^ _n ^ ^ _{J = 6 3}

3.3. (2H, d, 2-H), 5.03 (1H, d, J = 4.5Hz, 6-H),

5.20 (1H, s, CH-CO), 5.1-5.8 (1K, m, CH = C), 5.63 (1H, d, J = 4.5Hz, 7-H), 5.92 (1H, d, J = 12Hz, CH = C), 7.4 (5H, s, phenyl-H).

25 Example 28

D (-) - 2- (t-Butoxycarbonylamino) -2- (3-chloro-4-hydroxyphenyl ) -acetic acid (Compound 28)

A mixture of 6 g (0.03 mol) of 3-chloro-4-hydroxyphenylglycine and 9.8 g (0.045 mol) of di-t-butyl dicarbonate dissolved in 120 ml of 50% aqueous tetrahydrofuran (THF) containing 10 ml ( 0.071 mol) of triamine is dissolved for 3 h at room temperature. The mixture is concentrated to 60 ml and the concentrate is washed with ether. The

M / 24 279

~ 64 ·

aqueous layer is acidified with 6% hydrochloric acid and extracted with 200 ml of ether. The extract is washed with water and saturated NaCl solution, dried over MgS 2 O and evaporated to dryness to give 10 g of an oily residue which does not solidify on trituration with ethane-n-hexane.

Benzhydryl-7β- [D-2- (t-butoxycarbonylamino) -2- (3-chloro-4-hydroxyphenyl) -acetamido] -S-chloroethyl-cephem ^ -carboxylate (Compound 29).

CHCONH-NH

To a solution of 6.2 g (0.015 mol) of compound 2 and 5.4 g (0.018 mol) of compound 28 in 150 ml of dry THF are added 3 _S 7 g (0.018 mol) of DCC and the mixture is stirred for 1 h at room temperature. The precipitated during the stirring Dieyelohexy! Urea is filtered off, the filtrate is evaporated to dryness. The residue is extracted with 200 ml of ethyl acetate, and the extract is then washed with aqueous NaHCO3 solution, water and saturated IJaCl solution and dried over MgSO4. The filtrate is evaporated to dryness and the resulting oily residue is chromatographed on a silica gel column (Wako gel C-200 | 140g) eluting with toluene-ethyl acetate (10si). The desired fractions are collected and evaporated to dryness, yielding 10 g of the product 29 receives »

M / 24 279 -65-

1 ·

IR: ^ i ^ t 'cm "* ¹ 1790, 1720, 1630, 1500, 1370, 1240, 116O.

¹ ί

Example 30

Benzhydryl-7β- [D-2- (t-butoxycarbony! Amino) -2- (3-chloro-4-hydroxyphenyl) -acetamido] -3- (triphenylphosphino) -methylr. ^ -cephem-4-carboxylate-iodide (Compound 30) 10

CHCONH

NH · ν ν . _O

- CH ₂ P (C ₆ H ₅ ) ₃ J ^ö

To a solution of 10 g (0.0143 mol) of compound 29 in 10 ml of acetone is added 11.2 g (0.075 mol) of sodium iodide and the mixture is stirred for 30 min at room temperature. The mixture is concentrated to 30 ml. To the concentrate 200 ml of ethyl acetate are added and the mixture is washed with aqueous Na _p SpO, solution, water and saturated NaCl solution and dried over MgSO ^. The ethyl acetate solution is filtered and the filtrate concentrated to half the volume. 3.9 g (0.015 mol) of triphenylphosphine are added to the concentrate, and the mixture is stirred for 2 hours at room temperature. 300 ml of ether are added to the solution, whereupon a precipitate separates out, which is filtered off and dried. You get

30 9 f.2 g of phosphonoiodide 30.

IR '" Z cm 1780, 1680, 1490, 1350, 1240, 1150.

M / 24 279 -66-

1 , . ..

Benzhydryl 7ß- [D-2- (t-butoxycarbanyliunino) -2- (3-chloro-4-hydroxyphenyl) acetamido] -3 - [(Z) -1-propen-1-yl) -3-ceph

31) ''

HO-Kv, V-CHCONH 10

_ ,, *, v ^ ^ CH = CHCH ,; (Z)

A solution of 9 »5 g (9 mmol) of compound 30 in 200 ml of chloroform is washed with. a mixture of water (100 ml) and N NaOH (10 ml) and the mixture shaken for 3 min. The organic layer is washed with water and a saturated NaCl solution, dried over MgSO4 and concentrated to about half the volume. 20 ml of 90% acetaldehyde are added to the concentrate and the mixture is stirred at room temperature for 3 hours, treated with anhydrous MgSO4 and filtered. The filtrate is evaporated to dryness and the residue is chromatographed on silica gel 60 (Merck, 120 g) eluting with toluene-ethyl acetate (4si). The total> desired fractions are collected and evaporated to dryness. The residue is triturated with a mixture of ether, isopropyl ether and η-hexane to give 1 * 33 g of the blocked product 31.

IRs JP5 cm "" ¹ 1770, 1700, 166Ο ·, 1480, 1350, 1210, 1150.

Example 32

7β- [D-2-amino-2- (3-chloro-4-hydroxyphenyl) -acetamido] -3-C (Z) -1-propen-1-yl) -3-cephem-4-carboxylic acid (Compound '52 _t BMY-28060) ¹ V

M / 24 279

H = CHCE.

A mixture of 1.33 g (1.93 mmol) of compound 31 and 3 ml of trifluoroacetic acid is stirred for 30 minutes at room temperature. The mixture is diluted with 50 mL of ether isopropyl ether (1: 1) to give 1.072 g of the crude trifluoroacetate of compound 32, which is chromatographed on a prepPAK-C! _Q (Waters) filled column (80 mL) becomes. The column is eluted with water and 10bigem methanol. The eluate with 10bigem methanol is concentrated to a volume of 10 ml, whereby a crystalline precipitate separates, which is filtered off and washed with acetone and dried in vacuo over Pp ⁰ K. This gives 238 mg of the compound 32 (95% purity), mp. 180-185 ⁰ C (gradually increasing). The filtrate is concentrated to 5 ml and lyophilized to give 154 g of a second fraction which, determined by KPLC,. has a purity of 80 #.

cm " ¹ 1760, 1680, 1570, 1410, 1390, 1350,

IDcLX.

1290, 1270

^{(pH 7)} nm U) 232 (10000), 280 (10500)

NMR: i ^D 2 ^{0 + NaHCO} 3 ppm 1.68 (3H, d, J = ^6Hz , C = C-CH ₃ ), 3.25 (1H, d, J = 18Hz, 2-H), 3 , 57 (1H, d, J = 18 Hz, 2-H); 4.90 (1H, s, .CH-CO), 5.18 (1H, d, J = 4.5 Hz, 6-H), 5.72 (1H, d, J = 4.5 Hz, 7 -H), 5.5-5.9 (1H,

M / 24 279 -68-

, CH = C), 5.97 (1H, d, J = 12.Hz, CH = C), 7.02 (1H ₉ d, J = 8'Hz, phenyl-H), 7.30 (1H , dd, J = 8u.1.5Hz, phenyl-H), 7.50 (1H, d, J = 1.5Hz, phenyl-H).

1 _L e 1 33

D (-) -2- (t-Butoxycarbonylarnino) -2- (3,4-dihydroxyphenyl) -acetic acid (compound 33a) in admixture with its lrJ3jgid_j! TlrMP ^ ^ 33b)

A mixture of 3.66 g (0.02 mol) of 3,4-dihydroxyphenylglycine and 9.24 g (0.04 mol) of di-t-butyl dicarbonate dissolved in 120 ml of 50% aqueous THF solution containing 10 ml (0.071 mol) of triethylamine is stirred for 16 h at room temperature. The mixture is concentrated to 60 ml and the concentrate is washed with 100 ml of ether, acidified with N hydrochloric acid and extracted with ether (100.times.2 ml). The combined extracts were washed with water and saturated NaCl solution, dried over MgSO.sub.2 ₊ and evaporated to dryness to give 8 g of an oily residue which is a mixture of the desired 3,4-dihydroxyphenyl derivative and the 3- and 4-mono-O-BOC protected derivatives (BOC,

25 is t-butoxycarbonyl).

In game 34

Benzhydryl-7β- [D (-) - 2- (t-butoxycarbonylamino) -2- (3,4-dihydroxyphenyl) -acetamido] -3-chloromethyl-3-cephem-4-carboxylate (Compound 34a) in admixture with the 3- (and 4) -mno-0- _i- butoxycarbonyl derivatives (compound 34b)

M / 24 279 -69-

HO

A mixture of 8 g (0.0193 mol) of compound 2.8 g of the mixture according to Example 33 and 4.12 g (0.02 mol) of DCC in 200 ml of dry THF is stirred for 1 h at room temperature. The reaction mixture is evaporated to dryness. The residue is dissolved in 200 ml of ethyl acetate and filtered insoluble material (dicyclohexylurea) from. The filtrate is washed with aqueous NaHCO 3 solution, water and brine, dried over MgSO 4 and evaporated to dryness under reduced pressure. The oily residue is chromatographed on a silica gel column (Kieselgel 60, 130 g), eluting with toluene-ethyl acetate (5: 1) and toluene-ethyl acetate (2: 1). The eluate with toluene-ethyl acetate (5: 1) is collected and evaporated to dryness to give 9.5 g of a mixture of mono-O-BOC-N-BOC-dl protected derivatives (34b). The eluate with toluene-ethyl acetate (2: 1) is collected and evaporated to dryness to give 3 g of the 3,4-dihydroxyphenyl derivative (34a).

Compound 34a 30

I ^R: v? £ cm " ¹ 1770, 1720, 1690, 1500, 1370, 1240, 1150

NMR: cS ^CDC1 3 pp _m 1.42 (9H, s, C-CH _3), 3.4 (2H, br.s, 2H), 4.30 (2H, br.s, CH ₂ -Cl ), 4.85 (1H, d, J = 4.5Hz, 6-H), 5.07 (1H, d, J = 6Hz, CH-NH), 5.74 (1H, dd,

M / 24 279 -70-

1

J = 9 u.4.5 Hz, 7-H), 6.6-6.9 (3H, m, phenyl-H), 6.93 (1H, s, CHPh), 7.3 (10H, s , Phenyl-H).

5. Mixture ^

5! ^c ">" ¹ 1770, 1720, 1690, 1500, 1370, 1240, 1150

NMR; a ^CDC1 3 ppm 1.42 (9H, s, C-CH _3), 1.55 (9H _f s, C-CH _3), 3.4 (2H, br.s, 2H), 4 35 (2H, br s, CH ₂ -Cl), 6.9-7.1 (4H, ra, CHPh and phenyl-H), 7.3 (IOH, s, phenyl-H).

15 Example 35

Benzhydryl-7β- [D (-) - 2- (t-butoxycarbonylamino) -2- (3,4-dihydroxyphenyl) -acetamido] -3-triphenylphosphonomethyl- 3-cephem-4-carboxylate-iodide (Compound 35a)

^v CHCONH- τ- -

NH

₃ ) ₃ CO _. CH (C ₆ H ₅ ) ₂

A mixture of 3 g (4.4 mmol) of compound 34a and 3.3 g (22 mmol) of sodium iodide in 50 ml of acetone is stirred for 30 minutes at room temperature. The mixture is concentrated to dryness. The residue is extracted with 100 ml of ethyl acetate and the extract is washed with aqueous NapSpO ^ solution, water and saturated NaCl solution. After drying over MgSO 4, the extract is concentrated to 60 ml. To the concentrate is added 1.4 g (5.3 mmol) of triphenyl phosphine and the mixture is stirred for 1 h at room temperature. To the mixture is added 100 ml of ether, a low

M / 24 279

precipitate, which is filtered off and washed with ether. 3.2 g (70%) of the phosphonium iodide (35a) are obtained.

^IR: ^ max ^cm " ¹ 1780, 1680, 1480, 1430/1360, 1240, 1150.

10 15

Similarly, 9.5 g (12 mmol) of the mixture of the mono-O-BOC protected derivatives (34b) are allowed to react with sodium iodide and then triphenylphosphite to give 10.7 g (77%) of a mixture of the corresponding monomers -O-BOC-N-BOC-triphenylphosphoniomethyl derivatives (35b).

cm " ¹ 1770, 1720, 1680, 1480, 1430, 1360, 1240, 1140.

Example Benzhydryl-7β- [D (-) - 2- (t-butoxycarbonylamino) -2- (3,4-dihydroxyphenyl) -acetamido] -3- [(Z) -1 -propene-1-yl] - 3-ceph em-4-carboxylate (Compound 36a)

25

HO

CHCONH NH

CO ₂ C (CH ₃ )

CH = CHCH ^ (Z) CO ₂ CH (C ₆ H ₅ ) ₂

30 35

To a stirred solution of 3.15 g (3 mmol) of compound 35a and 10 ml of acetaldehyde in 50 ml of chloroform is added dropwise with stirring for 10 min 8 ml (4 mmol) of 0.5N sodium hydroxide and stirred, the mixture for 1 h at room temperature. The reaction mixture is washed with water and saturated NaCl solution, dried over MgSO.sub.2 ₊ .

M / 24 279 -72-

net and evaporated at reduced pressure. Chromatograph the oily residue on a silica gel column (Wako gel C-200, 60 g) eluting with chloroform (2 L) and 2% methanol in chloroform, under TLC control (chloroform / methanol = 10: 1). The desired fractions of the eluate with 2% methanol are collected and evaporated to dryness to give 0.8 g (40%) of the propenyl derivative 36a.

WMi $ ^CT3C1 3 ppm 1.28 (3H, d, J = GHz, C-CH _3), 1.42 (9H, s, C-CH _3), 3.25 (2H, s, 2H), 4.92 (1H, d, J = 4.5Hz, 6-H), 5.08 (1H, d, J = 6Hz, CH-NH), 5.3-5.8 (1H, m, CH = C), 5.80 (1H, d, J = 4.5 Hz,

7-H), 6.04 (1H, d, J = 11Hz, CH = C), 6.70 (2H, s, phenyl-H), 6.82 (1H, S, phenyl-H), 6 , 92 (1H, s, CHPh), 7.3 (1OH, s, phenyl-H).

Similar to the procedure described above, 10.5 g (9.3 mmol) of the mixture of 3- and 4-O-BOC-N-BOC-digested derivatives 35b are reacted with acetaldehyde to give 3.3 g (46%). of the corresponding 3-propentyl derivative 36b.

IRs ^ Sv ^cm " ^{1 1} ^ 70, 1700, 1500, 1370, 1240, 1150

NMR: I ^CDCl 3 ppm 1.4 (9H, s, C-CH ₃ ), 1.55 (9H, s, C-CH ₃ ),

3.25 (2H, s, 2-H), 6.07 (1H, d, J = 11Hz, CH = C), 6.9-7.1 (4H, m, CH-Ph, and phenyl-H ), 7,3-7,5

(1OH, m, phenyl-H).

M / 24 279

10

15202530

Example 37

7B- [D (-) - 2-Amino-2- (3,4-dihydroxyphenyl) -acetamido] -3 - [(Z) -1-propen-1-yl] -3-cephem-4-carboxylic acid (Compound 37, BMY-28068) '

H = CHCH.

(Z)

A mixture of 0.8 g (1.2 mmol) of compound 36a, 0.8 ml, anisole and 3 ml of trifluoroacetic acid is stirred for 5 min at room temperature and diluted with 25 ml of ether and 25 ml of isopropyl ether. The resulting precipitate is filtered off and washed with isopropyl ether to give 557 mg of the crude trifluoroacetate salt of the compound. A solution of the crude product in 10 ml of water is purified by column chromatography using 100 ml of a pad of PrOpPAK-C ₁₈ (Waters). The column is eluted successively with water and 5% methanol. The eluate with 50% methanol containing the desired product is concentrated to 5 ml and lyophilized to give 231 mg (47%) of Compound 37 (in zwitterionic form, 90% pure), m.p. Zers.).

^IR: ° max ^cm

-1

1760, 1690, 1580, 1530, 1400, 1360,

1290, 1270

UV: χ Phosphate buffer (pH 7) ^ _{(£ )} 233 (9200), 281 (11000)

Max

35

H / 24 279 -74-

NHR: (S ^D 2 ° ppm 1.68 (3H, d, J = 6Hz, C-CH ₃ ), 3.26 (1H, d,: J = 18Hz, 2-H), 3.58 ( 1H, d, J = 18 Hz, 2-H),

5.18 (1H, s, CHNH), 5.22 (1H, d, J = 4.5Hz, 6-H), 5.5-5.9 (2H, m, CH = C, and 7- H), 5.97 (1H, d, J = 11Hz, CH = C), 7-o5 (3H, m, phenyl-H).

Similarly, 3.3 g (4.3 mmol) of N, O-Dit-BOC protected derivative mixture 36b give 1.3 g (75%) of Compound 37 in zwitterionic form (90% purity), whose spectral data are identical with the data given above.

15 Example 38

D (-) - 2- (t-Butoxycarbonylamino) -2- (4-hydroxy-3-methoxy- phenyl) -acetic acid (Compound 38)

20 .. HO (;> -CHCO ₂ H

CO ₂ C (CH ₃ ) ₃

A mixture of 2.96 g (0.015 mol) of D (-) - 2-amino-2- (4-hydroxy-3-methoxyphenyl) -acetic acid and 3.6 g (0.0165 mol) of di-t-butyl dicarbonate in 100 ml of 50% aqueous THF containing 4.2 ml (0.03 mol) of triethylamine is stirred for 16 h at room temperature and the reaction mixture is concentrated to 50 ml. The concentrate is washed with 50 ml of ether, acidified with N hydrochloric acid and extracted twice with ether (2 × 100 ml). The combined extracts are washed with water and saturated NaCl solution. The dried extracts are concentrated to dryness to give 4.38 g of compound 38 as a foamy solid.

M / 24 279 -75-

NMR: <ί ^CDCl 3 ppm .1.4 (9H, S ₁ -C -CH-), 3.8 (3H, s, OCH ₅ ),

5.15 (1H, d, J = 6Hz, CH-NH), 6.85 (3H, s, phenyl-H).

Example 39

Benzhydryl-7β- [D (-) -2- (tert -butoxycarbonyloxy) -2- (4-hydroxy-3-methoxyphenyl) -acetamido] -3-chloromethyl-3- cephem-4-carboxylate ( Connection 39)

-CHCONH

f ^"1 - ^ - N. ^ ^N Cl -CH ₂ CO ₂ CH (C ₆ H _{5) 2}

A mixture of 4.3 g of Compound 38, 5 g (0.012 mol) of Compound 2 and 3 g (0.015 mol) of DCC in 150 mL of dry THF is stirred for 2 h at room temperature. The precipitated urea is filtered off, the filtrate is concentrated to dryness. A solution of the residue in 200 ml of ethyl acetate is washed with aqueous NaHCO 3 solution, water and saturated NaCl solution, dried over MgSO 4 and evaporated to dryness. The oily residue is chromatographed on a silica gel column (Kieselgel 60, 100 g), eluting with toluene-ethyl acetate (4: 1), under TLC control (toluene-ethyl acetate (1: 1) or chloroform-methanol (50: 1) : 1)] elutes. The desired fractions are collected and concentrated to dryness to give 7 g of the desired 3-chloromethylcephem compound 39 as a foamy solid. , .

M / 24 279

NMR: S, ppm 1.4 (9H ₁ ¹ S, C-CH _3), 3/45 (2H, br.s, 2H), 3.83 (3H, s, OCH _3), 4.32 (2H, s, -CH ₂ Cl), 4.92 (1H, d, J = 4.5Hz, 6-H), 5.13 (1H, d, J = 6Hz / CH-NH), 5 , 65 (1H, d, J = 6 Hz, NH), 5.80 (1H, dd, J = 8 u.4.5 Hz, 7-H).,. 6.85 (3H, s, phenyl-K), 6.95 (1H, 9, CH-Ph), 7.2-7.5 (10H, m, phenyl-H).

Be i 's ρ ie .1 40

Benzhydryl-7β- [D (-) - 2- (t-butoxycarbonylamino) -2- (4-hydroxy-3-methoxyphenyl) acetamido] -3- triphenylphosphino-naphthyl ^ -cephem ^ -carboxylate-iodide (Compound 40 )

HO

CHCONH NH

H ^ CO

CO ₂ C (CH ₃ ) ₃ CO ₂ CH (C ₆ H ₅ ) ₂

A mixture of 7 g (0.01 mol) of compound 39 and 7.5 g (0.05 mol) of sodium iodide in 100 ml of acetone is stirred for 30 min at room temperature and evaporated to dryness one-five. A solution of the residue in 200 ml of ethyl acetate is washed with aqueous NapSpO 2 solution, water and saturated NaCl solution, dried over MgSO 4 and concentrated to 100 ml. 3.1 g (0.012 mol) of triphenylphosphine are added to the concentrate and the mixture is stirred for 1 h at room temperature. 100 ml of ether are then added to the reaction mixture and the precipitated solid is filtered off, washed with ether and dried to give 5.8 g of the triphenylphosphonium derivative 40. The ethereal filtrate is concentrated to 10 ml, and 300 ml of ether are added to the concentrate, 0.9 g of a

M / 24 279

second fraction of the product. The total yield is 6.7 g. ^

Example 41

Benzhydryl-70- [D (-) - 2- (t-butoxycarbonylamino) -2- (4-hydroxy-3-methoxyphenyl) -acetamido] -3 - [(Z) -I -propene-1- yl] - 3-Cephem-4-carboxylate (Compound 41)

CHCONH NH

CO ₂ C (CH ₃ ),

CH = CHCH, (Z) CO ₂ CH (C ₆ H ₅ ) ₂

To a stirred mixture of 5.8 g (5.5 mmol) of compound 40 and 10 ml of 90% acetaldehyde in 100 ml of chloroform is added dropwise with stirring for 25 min 11 ml ι (5.5 mmol) of 0.5N sodium hydroxide. The mixture is then stirred for 2 h at room temperature. The reaction mixture is washed with water, then with saturated NaCl solution, dried over MgSO 4 and concentrated to dryness. The oily residue was chromatographed on a silica gel column (Kieselgel 60; 130 g) _f eluting with a mixture of toluene and ethyl acetate [the micro, mixing ratio was changed stepwise; 4: 1 (1.3 l), 3: 1 (1.1 l), 2: 1 (1.0 l) eluted. The eluate is collected in 20 ml fractions. Fractions Nos. 26 to 59 are combined and concentrated to dryness to give 830 mg of the desired 3-propentyl derivative 41 as a foamy solid.

M / 24 279

NMR

CDCl

5 -

ppm 1.35 (3H, d, -CH-CH ₃₎ / 1.4 (9H, s, C-CH _3), 3.85 (3H, s, 0-CH _3), 6.07 (1H, d, J = 11 Hz, -CH = C).

For s ρ i e '1

42

7β- [D (-) - 2-Amino-2- (4-hydroxy-3-methoxyphenyl) -acetamido] -j - [(Z) -1-propen-1-yl] -3-cephem-4-carboxylic acid (Compound 42; ΒΜΥ-28Ό97) _______

CHCONH NIU

CH = CKCH

A mixture of 830 mg (1.2 mmol) of compound 41, 0.5 ml of anisole and 2 ml of trifluoroacetic acid is stirred for 5 min at room temperature. The mixture is then diluted with 30 ml of ether and 30 ml of isopropyl ether. The resulting precipitate is filtered off, washed with isopropyl ether and dried to give 437 mg of the crude trifluoroacetate of compound 42. The crude product is taken on a with 100 ml PTePPAK-C ₁ q (Waters). , random column, eluting with water and 5% methanol. The eluate with 5% methanol is concentrated to 5 ml and lyophilized to give 225 mg of compound 42 (in zwitterionic form, 90% pure), mp 176-18O ⁰ C (dec.) T.

IR: v> _Mn , cm " ¹ 1760/1690, 1590, 1530, 1400, 1360, 1280 ma ^ c

_{: / V} phosphate buffer (pH 7) ^ (^ 235 (10000), 280 (11000)

Max . , ,

M / 24 279 -79-

NMR: £ ^D 2 ° ppm 1.68 (3H, d, J = 6Hz, C-CH ₃ ), 3.25 (1H,

d, J = 18 Hz, 2-H), 3.57 (1H, d, J = 18 Hz, '2 H), 4.01 (3H, s, OCH _3), 5.10 (IH, s , CH-CO),

5.19 (1H, d, J = 4.5Hz, 6-H), 5.78 (1H, d, J = 4.5Hz, 7-H), 5.5-5.9 (1H , m, CH = C), 5.98 (1H, d, J = 11Hz, CH = C), 7.07 (2H, s, phenyl-H), 7.17 (1H, br.s, phenyl -H). 10

HPLC: retention time = 9.3 min (0.02 M acetate buffer (pH 4) containing 15% acetonitrile).

Example 43

Isolating compound 42 from rat urine,. Compound 37 was administered

Six male Wistar rats (400-600 g) were placed in metabolic cages following oral administration of Compound 37 at a dose of 100 mg / kg. The urine was collected over a period of 14 hours / The rats were supplied with food and water as usual during the experiment. The following table shows the collected in the different periods

25 amount of urine.

0-2 h 2-4 h 4-6 h 6-24 h Total

Urinvolu-

meri (ml) 18 19.5 13 42 92.5 -

The urine (about 90 ml) was adjusted to pH 3 with N hydrochloric acid and filtered to remove a precipitate. The filtrate was chromatographed on a column filled with 300 ml of HP-20, eluting with 2 liters of water and 2 liters of 30% methanol followed by chromatography by HPLC. Those

M / 24 279 · -80- '

  ι. , '.' ·

Fractions containing the bioactive constituents of the eluate with 30% methanol were collected, concentrated to 10 ml and lyophilized to give 390 r of a brown solid. A solution of this solid in 20 ml of water was chromatographed on a column filled with 200 ml prepPAK-C _iQ (Waters). Successive elution with water, 5% methanol and 10% methanol. The first half of the eluate with 5% methanol was concentrated to 5 ml and lyophilized to give 44 mg of Compound 37 (70% purity) containing urinary contaminants. The second half of the eluate with 5% methanol was concentrated to 5 ml and lyophilized to give 36 mg of product containing a mixture of compounds 37 and 42 and urine. Impurities was. The eluate with 10% methanol (about 600 ml) was concentrated to 5 ml and lyophilized to give 38 mg of compound 42 (70% pure by HPLC). This product was rechromatographed on a column as above (40 ml) eluting with water, 5% methanol and 10% methanol. The desired fractions eluted with 10% methanol were combined and concentrated to 5 ml and lyophilized to give 16 mg of compound 42, which had a purity of 90% by HPLC [0.02 M acetate buffer, (pH 4)]. Acetonitrile (85:15)]; Mp. 180 ⁰ C (allmähl.Zers.).

30

cm "" ¹ 1760, 1690, 1590, 1530, 1400, 1360, 1280

Phosphate buffer (pH 7) _{nm (£} ) 233 (8200), 280 (8800) max

M / 24 279 - -81-1

NMR: ^D 2 ° el ppm 1, 68 (3H, d, J = 6 Hz, -C-CH ₃₎ -, 3.26 (1H,

d, J = 18 Hz, 2-H), 3.58 (1H, d, J = 18 Hz, 2-H), 4.01 (3H, s, OCH ₃ ), 5.12 (1H, s, CH-CO),

'5.21 (1H, d, J = 4.5Hz, 6-H), 5.78 (1H, d, J = 4.5Hz, 7-H), 5.5-5.9 (HI , m, CH = C-), 5.98 (1H, d, J = 11Hz, CH = C-), 7.07 (2K, s, phenyl-H), 7 »17 (1H, br.s , Phenyl-H).

The structure of the metabolite was determined to be 7β- [D (-) - 2-amino-2- (4-hydroxy-3) by comparison (NMR, IR, UY, HPLC) with Compound 42 prepared according to Examples 38-42 -methoxyphenyl) acetamido] -3-C (Z) -I -propene-1-yl] -3-cephera-4-carboxylic acid.

Claims (9)

wherein a hydrogen atom, OP ", a lower alkoxy means group or a halo atom, 2 P and P are hydrogen atoms or usual, in cations used for cephalosporin chemistry are amino, carboxy and hydroxy groups, a hydrogen atom, OP or a lower alkoxy group, and a hydrogen atom, a C 1 -C 4 alkyl, C ₇ -C ₁₄ aralkyl, heterocyclothio, C 1 ^ -Alkyl, C, -C ₄ alkoxy-C ₁ -C.-alkyl group or the group AIkX means, wherein M / 24 280 Alk an alkylidene-oiKn alkylene radical with 1 to. 4 carbon atoms, and X represents a bromine, chlorine or iodine atom ' ^{J and} the pharmaceutically acceptable acid addition salts, metal salts, amine salts or esters thereof, jQ marked dadurc h, that he A) a halide of the general formulas: '3 ^l ₁₅ - QCH ₂ X or R CH ₂ X wherein X is Cl, Br or J, with a triarylphosphine m reaction-inert, an organic liquid carrier at 20 to 150 ⁰ C to a phosphonium salt, B) the phosphonium salt in a water-immiscible, liquid, organic solvent with an aqueous base to a Phosphoranylverbindung the general NEN formulas: QCH = PAr ₃ or R ³ CH = PAr ₃ gO, ' C) a compound ilv ι uligemeinen formula: QClI l'Ar, under anhydrous bedimjungen at -40 ⁰ C. to t50 ° C in the mentioned, with water M / 24 280 immiscible liquid organic solvent with a carbonyl compound of the general formula 'R ³ CHO or under the same reaction conditions Compound of the general formula: R CH = PAr ₃ with a carbonyl compound of the general formula: QCHO to react, where Q means a remainder of the Koι: HO- / Λ- CIlUONIl R " (0) (0) AcNH (0) Ii-N M / .24 280 -Jf- 20 means, η, R ¹ , P ^{1 have} meanings and η, R, P, P, P and R d i ι; above Ac is an acyl group usually present in cephalosporin compounds, and B is an alkylidene or aralkyl phosphate protecting group D) into the thus obtained 3-substituted-7-amino ceph-3-em compound, if necessary after Splitting off one or more of the protective 12 'groups P, P, P, Ac and B, the 7-acyl radical the general formula: wherein R and R have the meanings given above, introduced and E) the compound thus obtained, if desired, converted into the pharmaceutically acceptable salts or esters. 2. 'Process according to item 1, characterized in that gO' - one uses starting and intermediate compounds of the general formulas given in point 1, wherein n, R ¹ , R ² , R ³ , P ¹ , P ² and P meanings given have, .35 M / 24 280 -JET- I- where at least one of the substituents R, 2 3
R and R are different from hydrogen. 3. Method according to point 2, characterized by that starting and intermediate connections are used 1 2 3, where η is 0 and P, P and P Mean hydrogen atoms. ' 4. Method according to item 1 for the preparation Substituted vinyl cephalosporin compounds of general formula XV, wherein R is AIkX,. *. , ^and ₍ ^! the acid addition salts and metal salts of those compounds in which η is 0 and P, P / P is a hydrogen atom, characterized by the fact that « in step A a halide compound of the general formula R CH-X and in step C a carbonyl "3 compound of general formula R CHO use det, wherein each R is AIkX. 5. The method according to item 4, characterized in that one uses a halide or carbonyl compound, wherein Alk is a methylene group and on X signify a chlorine atom. M / 24 280 ^ 6. A method according to item 1 for the preparation of substituted Z-configured vinyl cephalosporin compounds on the exocyclic double bond of general formula XV, wherein R is a methyl group, and the pharmaceutically acceptable acid addition salts and the pharmaceutically acceptable metal salts IQ ^{s of} the compounds of the above formula in which η is 0 and P, P and P represent a hydrogen atom, characterized in that in step A, a halide compound of the formula CH ^ CH ₂ X and in step C, a carbonyl compound of the formula CH-.CHO used. 7. The method according to item 6, characterized in that one uses starting and intermediate compounds, wherein η is for 20 mean me. In which η is 0 and P, P and P are hydrogen atoms. ^v 8. The method of item 6, characterized in that one uses starting and intermediate compounds wherein η is 0 and at least one of the substituents P, P and P is a conventional protective group. 9. Method according to item 8, characterized in that starting and intermediate compounds are used, in which, when P and P are a protective group, these are independently selected from trityl, chloroacetyl, formyl, trichloroethoxycarbonyl, t-butoxycarbonyl and benzyloxycarbonyl and, when P is a protective group, this is selected from benzyl, p-methoxybenzyl, p- Nitrobenzyl, diphenylmethy1, t-butyl, and 2,2,2-trichloroethyl. . M / 24 280 10. The method according to any one of the preceding points, characterized in that the following compounds are obtained as the process product: a) 7β- [D-2-amino-2- (4-hydroxyphenyl) acetamido] -3 - [(Z) -1-propen-1-yl] -3-cephem-4-carboxylic acid, b) 7β- [D-2-amino-2-phenylacetamido] -3 - [(Z) -1-propen-1-yl] -3-cephem-4-carboxylic acid, c) 7β- [D-2-amino-2- (3-chloro-4-hydroxyphenyl) acetamido] -3 - [(Z) -i-propene-1-yl-3-cephem-4 ¹⁵ carboxylic acid, d) 7β- [D-2-amino-2- (3,4-dihydroxyphenyl) acetamido] -3- [(2) -i-propene-1-yl-3-cephem-4-carboxylic acid _f 20 e) 7β-I D-2-amino-2- (4-hydroxy-3-mothoxyphenyl) - acetamido) -3 - [(Z) -1-propen-1-yl-3-cephem-4-carboxylic acid, f) 7β ~ [Dr-2-amino-2- (4-hydroxy [ _J hL, 'iiyl) acetamido] -3 - [(Z) -3-thioro-1-propyl-1-yl] -3-cephem -4-carboxylic acid, g) diphenylmethyl 7β- [D-2- (t-butoxycarbonylamino) -2- (4-hydroxyphenyl) aceLani.ido] -3- ((Z) -3-chloro-1-propene-1-yl] -3-cephem 4-carboxylate, h) diphenylmethyl 7β- [D-2- (t-butoxycarbonylamino-2- (4-hydroxyphenyl) acetamido] -3-t-3-iodo-1-propen-1-yl-3-cephem-4-carboxylate,   fr} M / 24 280 i) diphenylmethyl 7β- [2- (1-butoxycarbonylamino) -2- (4-hydroxyphenyl) acetamido I-3 - [(Z) -1-propene-l-5 yl] ceph-3-em-4-carboxylate, j) 7β- [ D-2-amino-2- (4-hydroxyphenyl) acetamido] -3 - [(Z) -1-buten-1-yl] -3-cephern-4-carboxylic acid, k) 7β- [D-2-amino-2- (4-hydroxyphenyl (acetamido) -3- vinyl-3-cephem-4-carboxylic acid, 1) 7Q- [D-2-amino-2-I4-hydroxyphenyl) acetamido] -3- [(Z) -3-phenyl-1-propen-1-yl] -3-cephem-4-carbon-15 acid m) 7β- [D-2-Amino-2- (4-hydroxyphenyl) acetamido] - '3 - [(Z) 3- (1H-1,2,3-triazol-5-yl) thio-1-propene 1-yl] -3-cephem-4-carboxylic acid and η) 7β- [D-2-Amino-2- (4-hydroxyphenyl) acetamido [-3 - [(Z) -3-metoxy-1-propen-1-yl] -3-cephem-4-rcar Bonic acid.